tag:blogger.com,1999:blog-56233087949286205762024-03-19T03:41:56.915-07:00S-STEM Scholar Amanda Goodson's BlogAmanda Chapmanhttp://www.blogger.com/profile/16951554565276475785noreply@blogger.comBlogger21125tag:blogger.com,1999:blog-5623308794928620576.post-87903927954855632812015-04-13T01:59:00.001-07:002015-04-13T01:59:12.193-07:00UpdateBlood: I performed the DNA extraction on all samples and will run horizontal agarose gel to see if there is a significant enough amount to warrant PCR amplification.<br />
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Algae: I helped Tony harvest his algal growth. Algal water was collected into falvon tubes and centrifuged. The supernatant was drained and the algal sediment was collected using a micrpippette and deposited into a metal pan. The pan was placed in an oven set at 70 degrees F for 45 minutes. Dried algae was scraped out of pan and collected in a beaker. The dried algal powder was ground using mortar and pestal (to make sure the cell walls were broken) and added to DIH2O a 30 cc syringe was used to remove water from beaker and inverted for 48 hours. The syringe shows 2 phases, a water phase and an oil phase. Oil extraction complete. The fish are still alive and happy and were added to the aquaponics fish family.<br />
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Protein: I began a new protein extraction and will run PAGE gels as soon as I read a couple more protocols. This week has been really busy with papers to write and exams in Calc and Chem.<br />
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Fish: The aquaponic fish are still quite happy with their salad bar root buffet and the addition of the algae fish. Half of the water was removed and replaced with DIH2O. I will check the pH when I return to lab next week. The fish have eaten almost all of the roots so I am going to grow more in the other tank. I noticed that there were a couple of plants that had roots stuck in the holes. I thought I would be helpful and pull the roots through. I grabbed a container out of the tank and it slipped out of my hand. I spent the next 30 minutes cleaning it up. No plants were harmed in the making of this blog. <br />
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<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-83403385185928678652015-04-13T01:40:00.000-07:002015-04-13T01:40:09.915-07:00Update on blood, protein, algal infection, and fishBlood: A protocol has been created and now I am waiting on primers. I collected blood, scalp hair, eyebrow hair, fingernail, and spit from myself, my daughter, and a friend that wishes to remain anonymous. we will call him Mr. XY. I will need to extract the DNA from these samples and then run a gel to see if DNA is present. Hopefully there will be a significant amount and I can proceed to PCR amplification.<br />
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Protein: I loaded a duplicate PAGE gel to the one that contains bands with 2 different biomarkers. Lane 8 contains 5uL Biorad Precision Plus Protein Dual Xtra Standards. This will measure proteins that range from 250kDa to 2 kDa.<br />
Lane 1 contain 5uL of Fisher Bio Reagent. This measure proteins that range from 116 kDa to 14.4 kDa. <br />
Running buffer was prepared with 100 mL 10x tris/glycerine/SDS buffer to 100 mL DIH2O. Gel ran at 140 volts for 1 hour and 30 minutes. <br />
Neither biomarker banded. I am not sure why but there could be many explanations. Possibilities include improper technique or equipment malfunction. I will research protocols and try again.<br />
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Algla infection: I was curious as to whether the bacteria that killed the algae is harmful to fish. If it is not, then it could possibly be used to eradicate unwanted algal growth from fish tanks. Two feeder fish were purchased from a local fish store and placed in a bowl containing 60 mL of contaminated algal water and 1 L of DIH2O. The bacteria are present. They are cocci, gram positive with a spikey appearance. As this bacteria matures the spikes grow into fimbrae that detach and float in the water. I took a 20 mL sample of water and introduced 2 antibiotic discs (penicillin and clindimycin) to see if the bacteria were affected. 48 hours later there were no bacteria present and the fish seem to be happy. Antibiotic discs were added to the infected water. After 6 days I came into the lab to see new algae growing in the once infected tank. <br />
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Fish: The plants have matured and the roots are growing through the holes in the bottom of the container and into the water. I placed the new plants into the tank and the fish immediately started nibbling. The pH is still steady at 8. Josh suggested removing half of the water and replacing it with DIH2O. I will do this next week. Two more fish were added to the tank in hopes that their excrement will lower pH.<br />
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<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-90302045660091575262015-04-13T00:47:00.000-07:002015-04-13T00:47:06.195-07:00Protein Extraction:<br />
Through creative optimism and a little hard work, I received 2 biomarkers this week (Thank you Josh). One is from Fisher (measures protein MW of 116.0-14.4 kDa) and one from Biorad (measures protein MW of 250-2 kDa). My plan was to run these two biomarkers on a 4-20% Tris-HCL PAGE and then use it to measure the bands on the 4-20% Tris-HCL PAGE that I ran last week (2 identical gels). I loaded to 10 uL of Fisher into lane one and 10 uL of Biorad into lane 8. Running buffer was prepared using 100 mL 10x tris/glycerine/SDS buffer to 900 mL DI water. The gel ran at 100 volts for 2 hours. What resulted was each biomarker produced one band about 1/4 from the top of the gel. I am not sure why this happened. The electrophoresis tank did not make the usual high pitch noise that it does when it reaches maximum voltage but it stayed at a steady 100 volts and I could see continuous bubbles so I didn't really worry about it. There are so many possibilities aas to why this run was not successful (equipment malfunction, defective product, and technique are at the top of my list so far). I will research this more and run a few more gels next week. <br />
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Blood: I am preparing to do a DNA extraction in order to visualize the double genome for blood type that I have been told that I have. I am meeting with Anil today to brainstorm some possible ideas. <br />
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Fish: The fish are still happy but the pH is still very stable at 8. They are also dependent on fish food and I would like them to eventually obtain optimal nutrition from the roots of the plants in the tank. Jeremy suggested that they do not like the plants that are growing and did a bit of research on fish palate. He discovered that fish enjoy bok choy, kale, arugula, and swiss chard. I used the extra tank to plant these. we will see if the fish enjoy their new salad bar.<br />
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Algal infection:<br />
So after taking a water sample and staining it, I discovered that there is indeed bacteria present. They are cocci,Gram positive bacteria that look pretty scary under the microscope. I streaked a McKonkey plate to make sure that they are gram negative and I also streaked a MSA plate to see if they are halophiles given the salinity of the water, although this may have been a byproduct from the algal cell lysis. <br />
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<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-70664581815310684952015-03-13T16:14:00.000-07:002015-03-13T16:14:15.760-07:00The electron crowd experiment<!--[if gte mso 9]><xml>
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<br />
What a busy but incredible week this has been. I received a phone call from
a fellow intern Sunday night. His project revolves around a specific type of
algae that can be grown, undergo an oils extraction which can be used for
bio-fuel. As fascinating as this sounds I know very little about this concept.
He stated that the algae he was harvesting "died" and that the project
was a failure and that he was going to trash the water and start over. I
explained that there is no such thing as "failure" in science and
asked him to meet me in the lab Monday morning to discuss what else this could
mean. He agreed to meet and talk with me before trashing the water. As I stood
in front of his tanks that morning I saw cloudy water with a light green
hue, sediment at the bottom of the tank, and a lot of hard work that had
been done. The brainstorming began. I started asking myself a few
basic questions:<br />
1. Why did the algae die? <span style="color: #cc0000;">Something killed it or
the growing conditions were not optimal</span><br />
2. What is the sediment on the bottom of the tank? <span style="color: #cc0000;">Put
it on a slide and look under a microscope. </span><br />
3. What would kill the algae? <span style="color: #cc0000;">Most likely a
microbe, collect it, stain it, identify it. </span><br />
4. Was the original algal sample tested for microbial growth when it
arrived? <span style="color: #cc0000;">No</span><br />
5. If a microbe attacked the algae where did it come from? <span style="color: #cc0000;">The lab or where it was shipped from</span><br />
6. What can we do to prevent this from happening again? <span style="color: #cc0000;">Identify the cause and implement a procedure to restore
the tank. </span><br />
7. Can we restore the algal growth? <span style="color: #cc0000;">If we can
identify the cause of death and eliminate it, it is possible to restore the
algal growth. </span><br />
8. What can we learn from this? <span style="color: #cc0000;">Without a proper
protocol and constant monitoring, algae will die. Also there could be a benefit
of a microbe that kills algae. </span><br />
9. What are possible benefits of a microbe that eliminates algae? <span style="color: #cc0000;">A natural algaecide for fish tanks. </span><br />
<br />
<span style="color: black;">Now I was ready to meet my friend. When he arrived
my excitement at all the new possibilities was contagious. A new project had
begun, but there was a lot of work to be done and only two of us. Lucky
for us, a perfect storm entered (Jeremy) and the electron cloud was
formed. Identifying the contaminate of the water was the first order of
business. I collected water from the tank, the pure algal culture, and the
bucket with the rest of the algal water that Tony brought back from the algal
lab in Carlsbad, Ca. I placed each specimen in a centrifuge for 10 minutes at
16 RPM. I extracted the supernatant added more sample and centrifuged again. I
repeated this two more times until I has a sufficient pellet at the bottom of
the PCR tube. I placed a sample of the pellet on a microscope slide and viewed
each sample under the microscope. I visualized algae and other microbes. I
decided to do a quick gram stain which ensued in an argument between the three
of us about the color of our new friend. I sad the color was purple but the
boys were adamant that the color was pink (I was wrong). Because there was
indecision, Jeremy streaked a TSA plate (to isolate a colony), and a macconkey
plate (to see if it was indeed gram negative or gram positive). After 24 hours
in the incubator, there was no growth. Jeremy asked if it were possible that
the microbe was anaerobic. We decided to light a candle to see. Tony was busy
testing the water sources for salinity, pH, and buying fish (to see if the
bacterium would negatively impact them). He also learned how to successfully
perform a gram stain (perhaps it is possible to teach an old dog new
tricks). A specimen of contaminated water was obtained (100mL) and 1 disc
of clindamycin and 1 disc of penicillin was added to this water. 24 hours later
there was zero microbial growth. Two feeder goldfish were obtained and placed
in a bowl with a mixture of 30 mL of contaminated water with 1 liter of DI
water. Salinity was 896 PPM. After the fish were added, the salinity dropped to
696 PPM. the following afternoon an additional 30 mL of
contaminated water was added to the fish bowl. 48 hours later, salinity was
1620 PPM, 30 mL contaminated water was added today and salinity is currently
2090 PPM. When the fish were first added the salinity decreased. That was the
only time a decrease in salinity was observed. MSA plate was
streaked today with contaminated water to see if the bacteria is sensitive to
salt. </span><br />
<span style="color: black;">This is what we have learned:</span><br />
1. The bacterium are anaerobic, gram negative, cocci, that start as round
spheres with spike-like projections and mature into what appears to be
filaments that grow in a bunch and break away from one another and infiltrate
water. Many other tests need to be done before identification can be made. <br />
2. The algal water arrived and had a salinity of around 5 PPM. After
contamination the salinity was so high that the meter read "error".
Either the bacteria or the algal death increased the salinity in the water. <br />
3. A combination of clindamycin and penicillin killed the bacteria in an
isolated study. <br />
4. The bacteria does not appear to harm fish. <br />
This has been an amazing week. I have truly loved this experience with this
electron cloud. You boys are the best! I look forward to working with you both
when we return from holiday.<br />
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<br />
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-88680236359341065132015-03-05T21:07:00.004-08:002015-03-05T21:07:51.253-08:00Save The Fish!<!--[if gte mso 9]><xml>
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<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="List Continue 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="List Continue 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="List Continue 4"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="List Continue 5"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Message Header"/>
<w:LsdException Locked="false" Priority="11" QFormat="true" Name="Subtitle"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Salutation"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Date"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text First Indent"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text First Indent 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Note Heading"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text Indent 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Body Text Indent 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Block Text"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Hyperlink"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="FollowedHyperlink"/>
<w:LsdException Locked="false" Priority="22" QFormat="true" Name="Strong"/>
<w:LsdException Locked="false" Priority="20" QFormat="true" Name="Emphasis"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Document Map"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Plain Text"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="E-mail Signature"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Top of Form"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Bottom of Form"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Normal (Web)"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Acronym"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Address"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Cite"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Code"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Definition"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Keyboard"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Preformatted"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Sample"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Typewriter"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="HTML Variable"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Normal Table"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="annotation subject"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="No List"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Outline List 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Outline List 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Outline List 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Simple 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Simple 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Simple 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Classic 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Classic 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Classic 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Classic 4"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Colorful 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Colorful 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Colorful 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Columns 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Columns 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Columns 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Columns 4"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Columns 5"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 4"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 5"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 6"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 7"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Grid 8"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 4"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 5"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 6"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 7"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table List 8"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table 3D effects 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table 3D effects 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table 3D effects 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Contemporary"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Elegant"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Professional"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Subtle 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Subtle 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Web 1"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Web 2"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Web 3"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Balloon Text"/>
<w:LsdException Locked="false" Priority="39" Name="Table Grid"/>
<w:LsdException Locked="false" SemiHidden="true" UnhideWhenUsed="true"
Name="Table Theme"/>
<w:LsdException Locked="false" SemiHidden="true" Name="Placeholder Text"/>
<w:LsdException Locked="false" Priority="1" QFormat="true" Name="No Spacing"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading"/>
<w:LsdException Locked="false" Priority="61" Name="Light List"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 1"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 1"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 1"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 1"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 1"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 1"/>
<w:LsdException Locked="false" SemiHidden="true" Name="Revision"/>
<w:LsdException Locked="false" Priority="34" QFormat="true"
Name="List Paragraph"/>
<w:LsdException Locked="false" Priority="29" QFormat="true" Name="Quote"/>
<w:LsdException Locked="false" Priority="30" QFormat="true"
Name="Intense Quote"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 1"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 1"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 1"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 1"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 1"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 1"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 1"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 1"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 2"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 2"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 2"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 2"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 2"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 2"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 2"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 2"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 2"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 2"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 2"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 2"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 2"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 2"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 3"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 3"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 3"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 3"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 3"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 3"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 3"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 3"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 3"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 3"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 3"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 3"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 3"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 3"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 4"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 4"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 4"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 4"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 4"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 4"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 4"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 4"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 4"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 4"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 4"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 4"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 4"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 4"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 5"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 5"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 5"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 5"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 5"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 5"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 5"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 5"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 5"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 5"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 5"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 5"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 5"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 5"/>
<w:LsdException Locked="false" Priority="60" Name="Light Shading Accent 6"/>
<w:LsdException Locked="false" Priority="61" Name="Light List Accent 6"/>
<w:LsdException Locked="false" Priority="62" Name="Light Grid Accent 6"/>
<w:LsdException Locked="false" Priority="63" Name="Medium Shading 1 Accent 6"/>
<w:LsdException Locked="false" Priority="64" Name="Medium Shading 2 Accent 6"/>
<w:LsdException Locked="false" Priority="65" Name="Medium List 1 Accent 6"/>
<w:LsdException Locked="false" Priority="66" Name="Medium List 2 Accent 6"/>
<w:LsdException Locked="false" Priority="67" Name="Medium Grid 1 Accent 6"/>
<w:LsdException Locked="false" Priority="68" Name="Medium Grid 2 Accent 6"/>
<w:LsdException Locked="false" Priority="69" Name="Medium Grid 3 Accent 6"/>
<w:LsdException Locked="false" Priority="70" Name="Dark List Accent 6"/>
<w:LsdException Locked="false" Priority="71" Name="Colorful Shading Accent 6"/>
<w:LsdException Locked="false" Priority="72" Name="Colorful List Accent 6"/>
<w:LsdException Locked="false" Priority="73" Name="Colorful Grid Accent 6"/>
<w:LsdException Locked="false" Priority="19" QFormat="true"
Name="Subtle Emphasis"/>
<w:LsdException Locked="false" Priority="21" QFormat="true"
Name="Intense Emphasis"/>
<w:LsdException Locked="false" Priority="31" QFormat="true"
Name="Subtle Reference"/>
<w:LsdException Locked="false" Priority="32" QFormat="true"
Name="Intense Reference"/>
<w:LsdException Locked="false" Priority="33" QFormat="true" Name="Book Title"/>
<w:LsdException Locked="false" Priority="37" SemiHidden="true"
UnhideWhenUsed="true" Name="Bibliography"/>
<w:LsdException Locked="false" Priority="39" SemiHidden="true"
UnhideWhenUsed="true" QFormat="true" Name="TOC Heading"/>
<w:LsdException Locked="false" Priority="41" Name="Plain Table 1"/>
<w:LsdException Locked="false" Priority="42" Name="Plain Table 2"/>
<w:LsdException Locked="false" Priority="43" Name="Plain Table 3"/>
<w:LsdException Locked="false" Priority="44" Name="Plain Table 4"/>
<w:LsdException Locked="false" Priority="45" Name="Plain Table 5"/>
<w:LsdException Locked="false" Priority="40" Name="Grid Table Light"/>
<w:LsdException Locked="false" Priority="46" Name="Grid Table 1 Light"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark"/>
<w:LsdException Locked="false" Priority="51" Name="Grid Table 6 Colorful"/>
<w:LsdException Locked="false" Priority="52" Name="Grid Table 7 Colorful"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 1"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 1"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 1"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 1"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 1"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 1"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 1"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 2"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 2"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 2"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 2"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 2"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 2"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 2"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 3"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 3"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 3"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 3"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 3"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 3"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 3"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 4"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 4"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 4"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 4"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 4"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 4"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 4"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 5"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 5"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 5"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 5"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 5"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 5"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 5"/>
<w:LsdException Locked="false" Priority="46"
Name="Grid Table 1 Light Accent 6"/>
<w:LsdException Locked="false" Priority="47" Name="Grid Table 2 Accent 6"/>
<w:LsdException Locked="false" Priority="48" Name="Grid Table 3 Accent 6"/>
<w:LsdException Locked="false" Priority="49" Name="Grid Table 4 Accent 6"/>
<w:LsdException Locked="false" Priority="50" Name="Grid Table 5 Dark Accent 6"/>
<w:LsdException Locked="false" Priority="51"
Name="Grid Table 6 Colorful Accent 6"/>
<w:LsdException Locked="false" Priority="52"
Name="Grid Table 7 Colorful Accent 6"/>
<w:LsdException Locked="false" Priority="46" Name="List Table 1 Light"/>
<w:LsdException Locked="false" Priority="47" Name="List Table 2"/>
<w:LsdException Locked="false" Priority="48" Name="List Table 3"/>
<w:LsdException Locked="false" Priority="49" Name="List Table 4"/>
<w:LsdException Locked="false" Priority="50" Name="List Table 5 Dark"/>
<w:LsdException Locked="false" Priority="51" Name="List Table 6 Colorful"/>
<w:LsdException Locked="false" Priority="52" Name="List Table 7 Colorful"/>
<w:LsdException Locked="false" Priority="46"
Name="List Table 1 Light Accent 1"/>
<w:LsdException Locked="false" Priority="47" Name="List Table 2 Accent 1"/>
<w:LsdException Locked="false" Priority="48" Name="List Table 3 Accent 1"/>
<w:LsdException Locked="false" Priority="49" Name="List Table 4 Accent 1"/>
<w:LsdException Locked="false" Priority="50" Name="List Table 5 Dark Accent 1"/>
<w:LsdException Locked="false" Priority="51"
Name="List Table 6 Colorful Accent 1"/>
<w:LsdException Locked="false" Priority="52"
Name="List Table 7 Colorful Accent 1"/>
<w:LsdException Locked="false" Priority="46"
Name="List Table 1 Light Accent 2"/>
<w:LsdException Locked="false" Priority="47" Name="List Table 2 Accent 2"/>
<w:LsdException Locked="false" Priority="48" Name="List Table 3 Accent 2"/>
<w:LsdException Locked="false" Priority="49" Name="List Table 4 Accent 2"/>
<w:LsdException Locked="false" Priority="50" Name="List Table 5 Dark Accent 2"/>
<w:LsdException Locked="false" Priority="51"
Name="List Table 6 Colorful Accent 2"/>
<w:LsdException Locked="false" Priority="52"
Name="List Table 7 Colorful Accent 2"/>
<w:LsdException Locked="false" Priority="46"
Name="List Table 1 Light Accent 3"/>
<w:LsdException Locked="false" Priority="47" Name="List Table 2 Accent 3"/>
<w:LsdException Locked="false" Priority="48" Name="List Table 3 Accent 3"/>
<w:LsdException Locked="false" Priority="49" Name="List Table 4 Accent 3"/>
<w:LsdException Locked="false" Priority="50" Name="List Table 5 Dark Accent 3"/>
<w:LsdException Locked="false" Priority="51"
Name="List Table 6 Colorful Accent 3"/>
<w:LsdException Locked="false" Priority="52"
Name="List Table 7 Colorful Accent 3"/>
<w:LsdException Locked="false" Priority="46"
Name="List Table 1 Light Accent 4"/>
<w:LsdException Locked="false" Priority="47" Name="List Table 2 Accent 4"/>
<w:LsdException Locked="false" Priority="48" Name="List Table 3 Accent 4"/>
<w:LsdException Locked="false" Priority="49" Name="List Table 4 Accent 4"/>
<w:LsdException Locked="false" Priority="50" Name="List Table 5 Dark Accent 4"/>
<w:LsdException Locked="false" Priority="51"
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FISH:<br />
I decided to save the fish! I was sad to hear that nobody picked up Jenni's
aquaponics project from last semester. I decided to take it on because they are
the friendliest fish that I have ever seen. The water was really low and the
plants were not looking very healthy. The pH is around 8 which is not healthy
for the fish or the plants. The plants are happy with a pH around 7.5 and the
fish will die at a pH of about 6.5 so pH 7 seems to be the happy number. I
added water and waited 3 days to see if the pH would lower. It did not. Adding
more fish (ammonium) will lower the pH but I was not sure how many fish to add.
A second tank was set up to become a spare fish home. If it gets too acidic in
the aquaponic tank they will have a safe place to go. 5 new fish were purchased
and added to the spare tank. After acclimating, 2 of these fish were added to
the aquaponic tank. Hopefully this will bring optimal pH to all of my new
friends. <br />
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Unknownnoreply@blogger.com2tag:blogger.com,1999:blog-5623308794928620576.post-51749235464670831442015-03-05T21:03:00.002-08:002015-03-05T21:06:48.390-08:00So many thingsThis week has truly been interesting . I found out that it is possible for a person to have two different blood types. It happens most often with twins exchanging genetic material in vitro and is commonly called chimerism. After receiving a letter from United Blood Services after a recent donation I was made aware that I may be one of these people. The letter also stated that they are not sure which blood I could be given if I ever required a transfusion because according to the test I have A and O blood. . I also am not a twin so this information was even more confusing. I decided to delve more into this and began by typing my blood. The test showed little to no agglutination in any of the serums. I called my mother to try to help me fill in the blanks. She stated that a fetus was removed through a D&C 27 months after my birth. After much research I learned that chimerism can be the result from one twin absorbing the genetic material from a nonviable fraternal twin fetus. I explained the situation and she confirmed that when I was 5 I had a minor surgery and she was told that I had a rare blood type and could not receive a transfusion if the need arises. She also confirmed that I was a very selfish child so the possibility of me absorbing a twin is highly probable. I would like to test my DNA to see if this is truly the case. I also want to mix different blood types with mine on a microscope slide in order to see if there is anyone that is compatible with me. A few brave souls volunteered this week but so far none of the RBC's got along with mine. I am currently looking for primers in order to get a final answer out of my DNA. <br />
<br />
PROTEINS:<br />
With the help of Josh and his mad hot gun skills I was finally able to successfully pour an agarose vertical gel and it was really cool. I loaded to 50 mL syringes with smelly SDS agarose gel and loaded the vertical plate while Josh kept the glass warm with a heat gun. I am still trying to figure out the best way to place the gel and the proper voltage and running time for the vertical chamber before I perform an electrophoresis with this. I am also waiting on bio-markers before I run and electrophoresis with the denser PAGE gels.<br />
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<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-36963540034500814332015-03-05T20:31:00.002-08:002015-03-05T20:32:55.607-08:00It's all about the gels this week. After reading a journal article titled <u>SDS Agarose Gels for Analysis of Proteins</u> I discovered that they were using mini-casettes (8x8x0.1 cm) that were loaded with agarose gel. I have previously been using polyacrylamide mini-cassette gels but the concentration was not dense enough to capture the specific proteins that I am trying to capture. I had some of these mini-casettes in the refrigerator. I took them apart using the key (long flathead screwdriver), being very careful to only break the side seals in order to keep the seal at the bottom intact. I used liquid tape to secure the edges. 0.5 g of agarose was mixed with 50 mL of SDS on a hotplate. A 5 mL syringe was used to load gel. Unfortunately the liquid tape did not hold and the gel escaped enough to allow air bubbles into the cassette rendering it useless. <br />
I decided to try a horizontal gel to see what kind of results I got. SDS (yelloe in color) was used to pour this gel (whitish-clear in color) and the wells were loaded with brain,urine and blood. A bio-marker was loaded but will only register proteins >6. After 88 minutes of electrophoresis at 100 volts I noticed that the gel was changing color. The gel was changing from yellow to clear as the proteins migrated from the negative pole to the positive pole. The positive reservoir was also changing from clear to yellow. When electrophoresis began both wells were clear from the buffer solution. I cut this gel in half and stained half of it with comassie blue and half of it with fast blast. The comassie blue showed banding but again, the bio-marker was not adequate so I still can not calculate the molecular weight of these proteins. The fast blast was too dark to visualize any banding. <br />
I became extremely frustrated and decided to attempt another vertical gel. I was pleasantly surprised to see a package from Genscript waiting for me. Inside was PAGE mini-cassete gels with the concentration needed to capture the 4 kda proteins. Next I will run the PAGE and attempt to successfully pour an agarose vertical gel.<br />
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<tr><td style="text-align: center;"><a href="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEjBiiKPoWiqLJ7OqhF1XBv7YKR0iOcarRJjqmSEWsArKJdppNCOwjZ65XUpk2u7Ffcv7Lg3BIzOROvd78yo_DsWr4tPWoL4AjV5vp2t40OHyxayA6rCDOdDBIvMsNu0khDf9o5zMgEdWACA/s1600/comassie.jpg" imageanchor="1" style="clear: left; margin-bottom: 1em; margin-left: auto; margin-right: auto;"><img border="0" src="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEjBiiKPoWiqLJ7OqhF1XBv7YKR0iOcarRJjqmSEWsArKJdppNCOwjZ65XUpk2u7Ffcv7Lg3BIzOROvd78yo_DsWr4tPWoL4AjV5vp2t40OHyxayA6rCDOdDBIvMsNu0khDf9o5zMgEdWACA/s1600/comassie.jpg" height="320" width="239" /></a></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Agarose gel stained with comassie blue</td></tr>
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<tr><td class="tr-caption" style="text-align: center;">Brain storming</td></tr>
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<tr><td style="text-align: center;"><a href="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiLQWTsJ7L6PfdG0FStyYiReZklJFo7ww1F0DEnncVIvLLou5PSip5V-l_4xIvTb4WCklJ7wr4W364FTXujcZjQ3W9-BoPfOZPeWWBVz6KRkKWePsvtwYYJuUTP21PjtnWPN2ECCIvqww88/s1600/horizontal.jpg" imageanchor="1" style="margin-left: auto; margin-right: auto;"><img border="0" src="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiLQWTsJ7L6PfdG0FStyYiReZklJFo7ww1F0DEnncVIvLLou5PSip5V-l_4xIvTb4WCklJ7wr4W364FTXujcZjQ3W9-BoPfOZPeWWBVz6KRkKWePsvtwYYJuUTP21PjtnWPN2ECCIvqww88/s1600/horizontal.jpg" height="240" width="320" /></a></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Reservoir well turns yellow</td></tr>
</tbody></table>
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<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><a href="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiqh_4elWttPBkB85oswyn57McoHTfZINLzjezgz0xqPFYa8i5KPbhSn08amTmIy9gTIkkLi3TdpkU0lS1ljHe-fGn5AHMyRIW0sfd0wj9h_fZVLITSIrzg7WIy-zrTpSY_0SXCyQZGYIVr/s1600/mini+pour+test.jpg" imageanchor="1" style="margin-left: auto; margin-right: auto;"><img border="0" src="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiqh_4elWttPBkB85oswyn57McoHTfZINLzjezgz0xqPFYa8i5KPbhSn08amTmIy9gTIkkLi3TdpkU0lS1ljHe-fGn5AHMyRIW0sfd0wj9h_fZVLITSIrzg7WIy-zrTpSY_0SXCyQZGYIVr/s1600/mini+pour+test.jpg" height="240" width="320" /></a></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Agarose mini-cassette </td></tr>
</tbody></table>
<br />
<div class="separator" style="clear: both; text-align: center;">
<a href="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiydouTrQj9o-WyGTW0kWe8JgeV5kZDrmUNzSST4fzB66FIXvEnrnIO_XF5CLmwtFjVc_mRvA6a9KMW4yH64VeQEzb2QwRpHAl5Te_jXJ5z1QqAPlZA5TtOmuTMa0OccwEfCEByauxC_X3p/s1600/my+sanctuary.jpg" imageanchor="1" style="margin-left: 1em; margin-right: 1em;"><img border="0" src="https://blogger.googleusercontent.com/img/b/R29vZ2xl/AVvXsEiydouTrQj9o-WyGTW0kWe8JgeV5kZDrmUNzSST4fzB66FIXvEnrnIO_XF5CLmwtFjVc_mRvA6a9KMW4yH64VeQEzb2QwRpHAl5Te_jXJ5z1QqAPlZA5TtOmuTMa0OccwEfCEByauxC_X3p/s1600/my+sanctuary.jpg" height="240" width="320" /></a></div>
<br />
<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-11130782068641168072015-02-17T22:41:00.001-08:002015-02-17T22:57:55.614-08:00PAGE RacesWelcome to the PAGE races! I will give a quick introduction to all of our competitors before we begin our 1 hour journey through pores big and small. Ladies and gentleman please fasten your safety belts. We will be traveling at a comfortable 200 volts and do not anticipate losing cabin pressure, but in the event that we do....just kidding. Here are the samples and the lanes they were loaded in. <br />
Lanes: <br />
1. Brain (non-hematoma)<br />
2. Brain (non-hematoma)<br />
3. Brain (hematoma)<br />
4. Brain (hematoma)<br />
5. Blood (sheep heart artery/ new)<br />
6. Blood (sheep heart artery/new)<br />
7. Blood (sheep heart artery/old)<br />
8. Blood (non-sheep)<br />
9. Urine<br />
10. Bio-Marker<br />
<br />
And the winner is......I can't tell. The polyacrylamide gels that I am using are long enough for the tiny proteins that i am trying to extract. The protein that I am after is around 4,000 kDa. This gel only goes to 10,000 kDa. The good news is that the proteins are actually moving in the gel which they were not doing with the previous SDS buffer I was using. This makes the smell of this new stuff worth it. The bad news is that the proteins are still a streak at the end of the plate. We have vertical electrophoresis tanks here in the lab which I can try. The only problem is that the gels cannot be purchased and polyacrylamide in it's polymer form, according to C3PO can "cause a very unpleasant death". I quickly imagined a bugs death and decided to research another way to run these vertical gels. I found a study that successfully used a 3% agarose gel (the bigger the percentage the smaller the pore size). My first attempt at pouring these gels was not successful. 1.8 grams of agarose is dissolved into 60 mL buffer solution. The solution is then injected into 2 panes of glass (about 8X11"). However, when I injected the gel it started to coagulate halfway down the glass. Oh, I almost forgot, the pieces of glass are so close together, the only thing I could find that would fit in-between them is an 18 gage needle. I also grabbed a used mini-gel cassette and super glued it together in order, and filled it with my new agarose solution to see if it can be reused. My second attempt at puring the vertical included the plates clamped with a rubber seal in-between the glass and placed in a water bath. This was also not successful due to water breaching the seal and compromising the gel. I have a new idea for attempt #3 but I will not ruin the surprise now :)<br />
It was great to see you all in the lab today. Thank you Jeremy for all of your help this week (including all of your witty banter). <br />
Also Thank you to Josh who has been so amazingly patient and accommodating. I appreciate the time you spend helping me pour through journal articles and combining all of the brainstorming sessions.<br />
<img alt="Displaying photo 1.JPG" class="aLF-aPX-J1-J3" height="200" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b9b679ce05e79e&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ8A4DTPuzxhH9SfUeN79o9RI1Hfgma91DiwWLOoQde75enLwWiWyPfqNSQFATGezkTRsxUBnBjbE6NEJp4X8bjERKLGbd6lpqQsFX5BKO3m5-PYD2z63sr4v-8&ats=1424241513200&rm=14b9b679ce05e79e&zw&sz=w1332-h536" width="150" /> <img alt="Displaying photo 2.JPG" class="aLF-aPX-J1-J3" height="149" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b9b679ce05e79e&attid=0.3&disp=inline&safe=1&attbid=ANGjdJ8KKirvn4zEH6BLy73VNp4sOXTHeu94Rz6if3-bpaSme_TzKxyj8sIFHk8LlaQNIWNAp_PdJzzFJ7W5YUN-0RomJQbBJF1HfOa5nfaWLw9aXQntok0L71mEcnI&ats=1424241513200&rm=14b9b679ce05e79e&zw&sz=w1332-h536" width="200" /><img alt="Displaying photo 3.JPG" class="aLF-aPX-J1-J3" height="200" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b9b679ce05e79e&attid=0.5&disp=inline&safe=1&attbid=ANGjdJ8ZI5lgekn-GURdffsBU6__6N4wtYF-IOfO5V8Y5rfie0IegvWsyBEWA8RFwGNEz-_n2ykR_-ebig-4uKaeB-4NPFksnXJJAkWeyfcjn-p8DP8HJNtJZ7QrDFo&ats=1424241513201&rm=14b9b679ce05e79e&zw&sz=w1332-h536" width="150" /><img alt="Displaying photo 4.JPG" class="aLF-aPX-J1-J3" height="149" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b9b679ce05e79e&attid=0.7&disp=inline&safe=1&attbid=ANGjdJ_Y9_PP_pTsg0znHU37EF3swMfQxD5SVctO247p7ZAqt4zSnW9bEERi_-DQQVT0P8l9Vzb-EPlET3GANNUfdRzkUqXRQQeg9SaFqQYNgpLW-0Tah3tnziHBEWI&ats=1424241513201&rm=14b9b679ce05e79e&zw&sz=w1332-h536" width="200" /><img alt="Displaying photo 5.JPG" class="aLF-aPX-J1-J3" height="239" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b9b679ce05e79e&attid=0.9&disp=inline&safe=1&attbid=ANGjdJ_S_BJxoH839sbzyLmQN6BL3v6jUaW4yGLv9ggmOjo2BgQvNBx_TNLEkVhizKVeU3V6EfHFNrJQ1qvDj2E1f9QK8YvSQ11Ae4VKYtfhb48ys_UyVeto8MJxQZ0&ats=1424241513201&rm=14b9b679ce05e79e&zw&sz=w1332-h536" width="320" /><br />
<br />
<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-3131121297932823702015-02-12T22:29:00.000-08:002015-02-12T22:29:04.159-08:00ElectrophoresisI am so happy to be back in the lab again! I am seeing many new faces and would like to say "welcome" to all of the new interns this semester. Words cannot describe how happy it made me to see familiar faces (Josh, Germie, Matt, Paul, D) you guys have become my friends. Now, enough sentiment...<br />
I am excited to continue with my project from last semester and I was super busy this week. My goal for this week was to complete a protein extraction using PAGE (Polyacrylamide Gel Electrophoresis). The most common protocol uses a steel ball mill (which we do not have and which is very expensive) so I was forced to be creative and create an alternative method (thanks Matt and Josh for the brainstorming sessions). I used sheep brains and hearts, placing 1 gram into a eppendorf tube with 250 uL of SDS buffer. I threw in a ball bearing and placed on the vortex until pulverized. This worked amazingly. Samples were then frozen, placed in boiling water, and centrifuged. The supernatant was extracted and a loading dye was added. Finally I set up my tank and loaded the gel. I am happy to report that the extraction was a success! However, the specific proteins that I am trying to extract have an atomic mass of 4514.04 kDa and the mini-cassette does not measure that small. Thwarted again! After another brainstorm session I have decided to perform another extraction using a vertical electrophoresis system. I had to become familiar with this contraption because I have never used one before and it looked a bit intimidating. Also, I will have to pour my own gels for this one and I was using polyacrylamide (which was pre-made and sent to the lab) which can be fatal in powder form. This didn't sound like a great idea to play with so I am going to try out a agarose gel. Monday I will make the cassettes and Tuesday I will load them and run them. Hopefully it will be a success!<br />
<img alt="Displaying photo 2.JPG" class="aLF-aPX-J1-J3" height="200" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14b37094c5cf93f0&attid=0.3&disp=inline&safe=1&attbid=ANGjdJ9xSVR220VyCXH1hO7zNzTMcckVL1s07jzDIWmHIDYDrlKO_UhmqIzu7jb2rrwgHCMpGW1DzHvWT0gHv4QjwdbxH4aXrYSKUceA6Tav-3Y4uzsmI5d-oyBoiCo&ats=1423807941097&rm=14b37094c5cf93f0&zw&sz=w1332-h537" width="150" /> <br />
<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-58353518962035082752015-02-12T22:25:00.003-08:002015-02-12T22:25:54.438-08:00New semester new protocol<div class="separator" style="clear: both; text-align: center;">
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It is great to be back in the lab. This semester I will
continue my exploration of the Beta-Amyloid proteins and antibodies associated
with Alzheimer's Disease. I was hoping to be able to obtain these proteins
and/or antibodies but they are extremely expensive. The protocol that I used
last semester extracted proteins from fish, but did not extract proteins from
the brains or blood. So my first step was finding a protocol for protein
extraction from brain tissue. Once again, the solutions needed for the extraction
protocol that appeared to yield the highest results are very expensive. I have
combined a few different protocols and with the help of Matt and Josh I am
ready to begin this journey. My first challenge came when my protocol called
for a pH 6.8 Tris HCl solution and the Trisma base we have in the lab has
a pH of around 9.8. With the help of a pH meter (which needed to be calibrated)
and some HCl I achieved my first mission. Next I needed to make an SDS buffer
which was simple to mix but one of the smelliest things I have ever been
exposed to. Beta- mercaptoethanol is the worst smelling compound that I have
ever smelled (the rest of the lab will agree, love you guys). The next step of this protocol was to pulverize the brain tissue in a ball mill grinder (which, again is very expensive). After a brief meeting with our local lab droid (C3PO) I decided to simulate a ball mill grinder. This meant placing 81 ball bearings (from the bearings on my roller skates) into a falcon tube along with the brain tissue and 250 uL of SDS buffer. The falcon tube was placed on the vortex for 8 minutes. Next week I will be preparing samples, loading PAGE gels and performing electrophoresis to see if this smelly protocol works.</div>
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<div class="MsoNormal">
</div>
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-66060009068540938642014-12-04T14:15:00.001-08:002014-12-04T14:15:37.959-08:00Presentation WeekWell the semester is coming to an end and I am not ready stop! This semester has been amazing. I learned so much working on this project. Although my final paper has been turned in I still have a few more tests that I would like to complete before this semester finally ends. I know know that proteins can be extracted from tissue samples, and that antibodies can be extracted from these proteins. I know that sheep brains can contain the beta amyloid proteins that are responsible for Alzheimers Disease (AD). Today I found a study that was performed at a university in Japan that was super empowering. They successfully measured the Beta amyloid proteins from individuals receiving treatment for AD. Now, all that is required is to put this all together and continue to work on my technique. Hopefully next semester will yield a tangible test that can be used in everyday application. However, my secondary hypothesis has proven to most challenging (Is it possible to consume an entire bag of tasty oh's in a semester?)<br />
<img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14a175e0e1b7b0d8&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ_ykeOrjicgUW6swsiUQqrUj2SpolMmB4uBhTv3aMorJvWJmvcRn-HWttvtN1fLE3iOggTK7KG9zrGnIrcjTWmzo_d6QiN0cGkJwujdFQJpZOCGj7twGa9ykXM&ats=1417731250763&rm=14a175e0e1b7b0d8&zw&sz=w1332-h536" /><br />
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<br />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-944014574049935062014-11-20T19:10:00.004-08:002014-11-20T19:19:33.303-08:00Just Keep Swimming!This week has been absolutely crazy for me. I had three exams and final project deadlines are quickly approaching! This week in lab I have worked on trying to come up with an idea for obtaining brain slices that are thin enough for me to see proteins but strong enough to survive the staining process. I started by selecting three brains that are positive for frontal lobe hematoma and three brains that do not have any visible hematoma. I am not sure that there is any significance to this but, I am curious as to why approximately 40 percent of the brains in an unopened bucket are positive and the rest are not. I then used a coring instrument to obtain three samples from each brain and placed them in the -80 degree freezer for 24 hours. The coring process was not much fun, the preservative smell and the close proximity needed for an accurate sample left me feeling a bit nauseous. Perhaps I will do only three at a time for my next coring session. Next I very slowly made slices using my makeshift microtome. I then took these slices and stained them using crystal violet. I have been able to slice them extremely thin but once I start staining them they disintegrate, littering the sink with brain matter which is a bit gross. The problem with such thin slices is that they thaw once they hit the room temperature microscope slides. Perhaps freezing them again before staining along with the microscope slides will help? I also got to spend time discussing projects with some of my S-STEM colleagues, celebrated Matt Hills birthday, and tried to offer a warm welcome to our newest STEMmers who were busy with their unknown microbe projects. All in all, it was a great week and I am excited to see everyone's presentations in the very near future!<br />
<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><img alt="Displaying IMG_2928.JPG" class="aLF-aPX-J1-J3" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=149d04dac6068388&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ8FoSU7ASokKwdq_LPnOw5wUZO5eDn_OqMPuk7jDVMXBJP4ZKZ0Hv9zdgH1fyDHF4PgqwsIqzmKllV5Gt9KJbi0pWDSJAUBqixnCdkKxl0YjTgjsVebxdfbAhQ&ats=1416539088886&rm=149d04dac6068388&zw&sz=w1332-h537" style="margin-left: auto; margin-right: auto;" /></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Hematoma Brain</td></tr>
</tbody></table>
<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><img alt="Displaying IMG_2931.JPG" class="aLF-aPX-J1-J3" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=149d04c74c838c1d&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ9l2LmqyxP3ozU6_CNCge8Wwahb3CRtCfQ5BVLrO0QiMMcVCwVG7ItHi7fuFmQiekBr84LngzsAKNZtm_MtJUiqkOurCFhWYNtek35D79apsxqkItqFHefj_ZI&ats=1416539088904&rm=149d04c74c838c1d&zw&sz=w1332-h537" style="margin-left: auto; margin-right: auto;" /></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Normal Brain</td></tr>
</tbody></table>
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-84882458534635400552014-11-13T15:12:00.003-08:002014-11-13T15:14:01.796-08:00S-STEM Field tripLast Friday I had the good fortune of my first S-STEM field trip. We went to Dreamy Draw park and I got to hike and experience geocaching for the very first time with some pretty amazing people. Thank you to our fearless (and extremely knowledgeable) tour guides Matt and Josh. I learned about femoral drainage and the mining of mercury that used to take place there. It was a much needed break from this stressful semester, and I am so happy that not only has S-STEM been an amazing experience, but I did not realize the bonds that would be formed from spending so much time together in the lab.<br />
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<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=149ab5fbc9600301&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ86Cdh8M4Y3riar1oT9d3Qeft-6rumQy_UkflC2NxxS0BOPll_0hmtmynfnuNCWLpH9DIeOvrPzxhsTMWlNDEV_wWtjyJ2kfDAagk3MryYl4KB-kC7niE77xNg&ats=1415919512236&rm=149ab5fbc9600301&zw&sz=w1332-h537" style="margin-left: auto; margin-right: auto;" /></td></tr>
<tr><td class="tr-caption" style="text-align: center;">This is a pic of Matt and I attempting microtome #2</td></tr>
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Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-54547790707375160312014-11-06T20:50:00.002-08:002014-11-06T20:50:56.024-08:00The search for a new brainThis week has been an exciting eek for me in the lab. I have successfully created a microtome! Ok, it is a very archaic design, but functional nonetheless. I used a 20 cc syringe, a brass corer (size 7), and a blade from a box cutter. The tip of the syringe was removed using a box cutter. After much measurement I realized that the diameter thickness that I wish to acieve just happens to be the distance between the cc markings on the syringe. Matt Hill used a saw to create an opening for my blade. the brain is cored and placed into the syringe. a smaller corer is used to plunge the brain down the syringe until it is pressed against the blade resting in its position within the cut in the syringe. when the blade is released the brain is moved ever so slowly forward and the blade slices through it. This little device has created some adequate samples for staining (using crystal violet) so I am very happy with it. However, the lego microtome is still in the process of being built (thanks to Matt Hill) but is not quite finished yet. The first brain that I sectioned was chosen due to the presence of a hematoma on the frontal lobe and a decreased frontal lobe size. This brain contained proteins when stained and viewed under the microscope. However, frozen brain tissue is much easier to cut so I placed it in the -80 degree freezer so that it would be super frozen. Well, when I applied the pressure to it in order to core it...it exploded into pieces which flew all over the bench and floor. although impressive it was really unpleasant to clean up (it defrosts quickly in such small pieces.) Now brainless, I picked another brain randomly from the bucket. after getting a sample under the microscope I quickly realized that it did not contain the proteins that I was looking for. I chose another brain randomly and again obtained negative results. I began searching the buckets for a third brain vowing to be picky this time to find one that shared the same characteristics as the first brain. I found the perfect specimen at the bottom of the bucket. Unfortunately I ended up dipping my leather banded watch in the process of retrieving it, which my fellow STEMmer Jeremy found extremely entertaining. This brain is positive for proteins. I am not sure if this is coincidence or not but it is defiantly interesting.<br />
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<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><img height="292" id="irc_mi" src="http://media.scbt.com/image.php?image=/i/26/30/263014.jpg" style="margin-left: auto; margin-right: auto; margin-top: 0px;" width="400" /></td></tr>
<tr><td class="tr-caption" style="text-align: center;">Beta amyloid present in brain tissue.</td></tr>
</tbody></table>
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http://www.google.com/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAYQjB0&url=http%3A%2F%2Fwww.scbt.com%2Fdatasheet-53822-beta-amyloid-20-1-antibody.html&ei=Kk9cVNP1MY72iQLCn4G4AQ&bvm=bv.79189006,d.cGE&psig=AFQjCNGoJW91fjtiLzgLVGozUWqAPsl6bA&ust=1415422087904595<br />
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-13705011441828019882014-10-30T16:51:00.001-07:002014-10-30T16:51:21.661-07:00Brains, Stains, and PainsSo this week was filled with good news and bad news. The bad news is that my western blot was not successful, however I did learn a ton from just completing the process. The good news is that I have completed collecting data for this part of my project and I am confident that I can extract the amyloid proteins should I be lucky enough to find a sample that contains them. I read an interesting article pertaining to these specific proteins being present in the brains of sheep. This is super lucky for me due to the abundance of them in our lab. I have read many articles related to staining proteins and which stains work best. It seems that; crystal violet, Iodine, and comaisse blue seen to be the most popular. I prepared brain samples on slides and used each stain just to see if one was easier to interpret than the others ( and because I was super curious). I liked the crystal violet best but my samples were too thick. A microtome would make this step so much easier but they are pretty expensive (although a lego version is currently under construction thanks to Matt H and Josh). I need each sample to be no thicker than 250 nm. Stains have been ordered, so next week promises much fun!<br />
<img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14963115af2c8cb4&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ9z6NQWrvS12Q9lY_gtRCRsVybRkiq2lWyBrnd6ARKjn1bFD61u68x4pmlwT0hWaQsLMcnCzL_blBOAKWTuCbwwP08tNCWQWLv6v18_ECaLoGTp2gkjf6Yvv1M&ats=1414713013012&rm=14963115af2c8cb4&zw&sz=w1332-h537" />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-25878824559273674212014-10-23T22:57:00.001-07:002014-10-23T22:58:00.207-07:00This Weeks Post is Brought to you by the Letter JJosh, J-E-L-L-O, jousting....three words that describe this week in the lab. Protein extraction through electrophoresis was not something that I was not familiar with before this semester. I successfully cut up my frozen fish samples into 0.25cm, watched C3PO add DDT to it (its harmful if inhaled), placed it in the water bath for 5 minutes at 95 degrees Celsius, loaded my gel (which comes pre-assembled in a very hard plastic cassette, added buffer to the tank, set the volts at 200, and crossed my fingers for about an hour. After the cycle was complete I rinsed the gel cassette and realized that this particular cassette requires a "key" which is not included, but can be purchased for $5.00 (plush shipping/handling). Let's Just say that I was not pleased with the manufacturers of this particular product. I had spent hours on this particular gel and was not going to lose my results because of a silly "key". I did some quick investigating of the mechanics of this cassette and decided that the most rational approach would be to just break it. I began by using the plastic comb that was removed from the wells of the gel but the cassette was not budging and was not giving up. Luckily Josh (the biker scout), sensing my frustration at Jousting with this cassette, came to the rescue with an extremely large flat head screwdriver. I quickly accepted this gift, had a fleeting thought of smashing the cassette with the tool, and finally decided to take his advice and use it to pry the thing apart. After a few minutes the cassette acquiesced and released my beloved gel. After it was rinsed and stained (thanks again to Josh for help with this process) I recorded my results. I was left with a gel full of data that was going to start to degrade which gave me a very sad feeling. I was not ready to part with this little masterpiece so I immediately went online to find out how others have successfully handled this situation. The answer was immediate and brilliant.....J-E-L-L-O! Gels can be preserved in a 90/10 Jello solution for long lengths of time. Now....on to Western Blotting!!<br />
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<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><br /></td></tr>
<tr><td class="tr-caption" style="text-align: center;"><br /></td><td class="tr-caption" style="text-align: center;"><img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" height="320" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=1493fdb69517e02f&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ9PT2GW8uGl3LWGTNJVj4rQYgfhUM1wo6ZUB1YB8SJFjsmwHRnHcDSRchF5b4FGsOSMS1T7NXcWuPFuLwsKKNKyMrsC8oKPzh4n7gHNSlqWFbj5TmGxH4QpQBQ&ats=1414129718399&rm=1493fdb69517e02f&zw&sz=w1332-h537" width="240" /><img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" height="320" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=1493fdb05090df70&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ8DiS03KzS44q-NwEJ7gI6qQbNtvcPnQ1uNgL1mxuv3mAcZVNzrCuJ9mE7magoLQAtlp5vcdXQjV6gHeZLqfh0Pnjt6MrTju5OawhBeP5rObsuzRDhr_jJpKBo&ats=1414129718413&rm=1493fdb05090df70&zw&sz=w1332-h537" width="240" /> </td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"> <img alt="Displaying photo.JPG" class="aLF-aPX-J1-J3" height="200" src="https://mail.google.com/mail/u/0/?ui=2&ik=2fd138127d&view=fimg&th=14940b3b204d8cca&attid=0.1&disp=inline&safe=1&attbid=ANGjdJ_TPsoZ5NDiiTAPyN9Bdud6jA5SQmwSbpKBnBMUYMwShsEcA8slLzy37LSLXhjIhOVtiMHzb10G38z-zM-wvpqvZNTPlxCX8H075UmcDybGo1oZ9WkJNhyeYM8&ats=1414129967250&rm=14940b3b204d8cca&zw&sz=w1332-h537" width="150" /></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"></td><td class="tr-caption" style="text-align: center;"><br /></td></tr>
</tbody></table>
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-61603846125341215262014-10-09T21:09:00.000-07:002014-10-09T21:10:36.718-07:00One Fish, Two Fish....This week I started the protein extraction process from five different
fish species. Biker Scout provided me with salmon and swai (which I have
never heard of) and we had a lively conversation of whether or not
canned tuna fish would work for this project (the answer was no, but we
both decided that it would be fun to try later). C3PO came to the rescue (with the help of the lab-mobile and some mechanics provided by our biker scout) and
provided very aromatic samples of catfish, dover, and halibut (which everyone else in the lab appreciated).. I am
very curious to find out how successful this technique is; for I would
like to try more evolved organisms after I master the fish. I also spent
some time swabbing a fellow stemmer which turned out to very amusing
and fun after the third hour. My fish have been cut up into 2.5 cm3
pieces and are marinating in buffer solution as I type this. My hope is that I will have results for my next post.<br />
<img height="412" id="irc_mi" src="http://www.glofish.com/files/GloFish-Fluorescent-Fish-Group-Photo_wPurple.jpg" style="margin-top: 0px;" width="618" /> Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-48323298176094743492014-09-30T16:09:00.000-07:002014-09-30T16:22:06.030-07:00Electrophoresis V.S Western Blotting for detection of a proteinSo after some intense dialog with the Biker Scout and C-3PO I think that I may be a bit closer to a project idea, even though it seems very far away from my goal upon first glance. This semester I will be using electrophoresis to isolate a specific protein. I will then use western blotting to introduce antibodies to the protein. Once I become fluent in this process my hope is to actually go after more specific human proteins/antibodies. I am curious to know if the antibodies produced from the beta amyloid protein could be used to create a test that would show the varying degree of beta amyloid proteins if they were present in an individual's blood. The first question that I had was; "Can beta-amyloid proteins be detected/isolated from urine and/or blood"? The answer is yes, they can be detected in blood, cerebral spinal fluid, and urine. My next question was,"Why is there not an inexpensive urinalysis (the least invasive procedure for the patient) that could detect the amount of beta amyloid proteins excreted through urine"? If the theory of the accumulation of these proteins or plaques is what causes the cognitive decline associated in Alzheimer's Disease is correct, than why are we not incorporating a device that will detect these plaques BEFORE they have time to build up? Many health care professionals already incorporate urinalysis into their annual physical protocol, why are we not monitoring the production of these proteins? Currently, the testing for these proteins is extremely costly and is almost always done on individuals that are ALREADY experiencing cognitive decline. I guess what I am asking is; when will we start to become proactive and get ahead of this disease, instead of ignoring it until it takes up space in our heads?<br />
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<img height="412" id="irc_mi" src="http://diverge.hunter.cuny.edu/%7Eweigang/Images/10-12_westernblot_1.jpg" style="margin-top: 0px;" width="548" /><br />
Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-40987782567039627132014-09-25T16:41:00.000-07:002014-09-30T14:59:43.929-07:00Change in Plan<br />
So apparently FISHing is super expensive, so I will not be doing it this semester. So with a heavy heart I will have to choose another project. I have always been curious about the beta amyloid plaques that accumulate in the brains of individuals affected with Alzheimer's Disease. These plaques are actually a sequence of 55 amino acids that most every person produces. The ability to excrete these proteins through urine reduces the probability of one being affected by this disease. When a person's body does not excrete the beta amyloid proteins they start to build up, like plaque on teeth. The once happy neurons frolicking happily in the brain are now plagued by these protein formations that prevent communication between one another. It takes many years for these formations to effect cognition which is why Alzheimer disease is usually reserved for the elderly population. I would like to see if I can isolate protein from blood in order to eventually acquire an antibody that I might study. If this project is successful, I would like to try to isolate beta amyloid protein from human blood. I wonder if I could start this process with blood from another organism? <br />
<img src="http://www.chemicalbook.com/CAS%5CGIF%5C131602-53-4.gif" height="364" id="irc_mi" style="margin-top: 24px;" width="363" />Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-79028792209541734762014-09-17T16:40:00.000-07:002014-09-18T10:23:19.380-07:00What is the Difference Between FISH and a Karyotype?FISH analysis will tell you if a specific chromosome is present and how many chromosomes there are and is much faster than a karyotype. FISH is used commonly for diagnosing down syndrome during pregnancy using amniotic fluid. Downs syndrome is diagnosed when there is a trisonomy on chromosome #21 (that is a number sign not a hash-tag). Now, what it will not tell you is the structural composition of the chromosome (like a karyotype) which is equally as important as knowing how many chromosomes are present. FISH analysis can take from 7-10 days. A karyotype can take up to two weeks to yield results. So, they are similar but very different procedures. Comparing FISHing to Karyotyping (not sure if either of those words are real words but you understand me for all intents and purposes) is like comparing an X-ray to an MRI. The X-ray is great for confirming the general location of pain that a patient is experiencing, but an MRI may be needed if there are further questions regarding structural integrity.So this semester I will learn to FISH, and if it is fun maybe I will try to karyotype (not myself though because I do not need to know that much about myself. It is already hard enough to compartmentalize the information I already have) totally kidding (kind of) :)<br />
<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="margin-left: auto; margin-right: auto; text-align: center;"><tbody>
<tr><td style="text-align: center;"><img height="412" id="irc_mi" src="http://www.breenlab.org/img/cytogenetics.jpg" style="margin-left: auto; margin-right: auto; margin-top: 0px;" width="313" /></td></tr>
<tr><td class="tr-caption" style="text-align: center;">FISH (property of Breen lab at NCSU)</td></tr>
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<table align="center" cellpadding="0" cellspacing="0" class="tr-caption-container" style="float: left; margin-right: 1em; text-align: left;"><tbody>
<tr><td style="text-align: center;"><a href="http://scigjt13.files.wordpress.com/2011/03/karyotype.gif" imageanchor="1" style="clear: left; margin-bottom: 1em; margin-left: auto; margin-right: auto;"><img border="0" height="276" id="irc_mi" src="http://scigjt13.files.wordpress.com/2011/03/karyotype.gif" style="margin-top: 68px;" width="300" /></a></td></tr>
<tr><td class="tr-caption" style="text-align: center;">KARYOTYPE</td><td class="tr-caption" style="text-align: center;"><br /></td><td class="tr-caption" style="text-align: center;"><br /></td></tr>
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Unknownnoreply@blogger.com0tag:blogger.com,1999:blog-5623308794928620576.post-75127631148475753862014-09-16T15:14:00.000-07:002014-09-16T15:14:56.369-07:00FISHing<blockquote class="tr_bq">
Hi there, this is my first ever blog so please keep that in mind while reading. I guess I will start with the first two questions that I had when I learned about FISH; what is FISH and why do I care? Fluorescence in situ hybridization (FISH) is a laboratory technique for
detecting and locating a specific DNA sequence on a chromosome. The
technique relies on exposing chromosomes to a small DNA sequence called a
probe that has a fluorescent molecule attached to it. The probe
sequence binds to its corresponding sequence on the chromosome.(<small>http://ghr.nlm.nih.gov/glossar y=fluorescentinsituhyb ridization)</small>. FISH is used by geneticists in order to diagnose abnormalities within chomosomes, which can save lives. That is why I care. I am a little nervous about attempting this project because it takes extreme precision and requires that measurements be conducted at specific times. I am super excited and ready to get started!</blockquote>
<a href="http://www.nature.com/scitable/topicpage/fluorescence-in-situ-hybridization-fish-327#"><img alt="This multi-panel diagram shows fluorescence in situ hybridization in five steps. The double-stranded target DNA is depicted in blue and the DNA probes are depicted as shorter red segments. In panel B, the probes are either directly or indirectly labeled with a fluorophore for visualization. Probes labeled indirectly with a hapten are shown with semi-circle protrusions on both strands. Probes labeled directly with a fluorophore are shown with full-circle protrusions on both strands. The target and probe DNA are denatured and hybridized in panels C and D, respectively. In panel C, the two strands of each DNA segment separate. In panel D, an indirectly-labeled probe is shown binding to the target DNA at left, while a directly-labeled probe is shown binding to the target DNA at right. The extra visualization step required for indirectly-labeled probes is shown in panel E." src="http://www.nature.com/scitable/content/35120/10.1038_nrg1692-f1_mid_1.jpg" title="" /></a><br />
<div class="imageCaption pipeblack bold nopad">
<a class="pipeblack bold" href="http://www.nature.com/scitable/topicpage/fluorescence-in-situ-hybridization-fish-327#"><img alt="View Full-Size Image" class="fleft borderNone" src="http://www.nature.com/scitable/natedimages/plus_box.gif" />Figure 1: Principles of fluorescence <i>in situ</i> hybridization (FISH).</a></div>
<div class="imageLegend pipeblack nopad">
<a class="pipeblack" href="http://www.nature.com/scitable/topicpage/fluorescence-in-situ-hybridization-fish-327#" style="font-weight: normal !important;">(a)
The basic elements of FISH are a DNA probe and a target sequence. (b)
Before hybridization, the DNA probe is labeled by various means, such as
nick translation, random primed labeling, and PCR. Two labeling
strategies are commonly used: indirect labeling (left panel) and direct
labeling (right panel). For indirect labeling, probes are labeled with
modified nucleotides that contain a hapten, whereas direct labeling uses
nucleotides that have been directly modified to contain a fluorophore.
(c) The labeled probe and the target DNA are denatured. (d) Combining
the denatured probe and target allows the annealing of complementary DNA
sequences. (e) If the probe has been labeled indirectly, an extra step
is required for visualization of the nonfluorescent hapten that uses an
enzymatic or immunological detection system. Whereas FISH is faster with
directly labeled probes, indirect labeling offers the advantage of
signal amplification by using several layers of antibodies, and it might
therefore produce a signal that is brighter compared with background
levels.</a></div>
© 2005 <a class="inlineLinks" href="http://www.nature.com/" target="_blank">Nature Publishing Group</a> Speicher, M. R. <i>et al.</i> The new cytogenetics: blurring the boundaries with molecular biology. <i>Nature Reviews Genetics</i> <b>6,</b> 784 (2005). All rights reserved.<blockquote class="tr_bq">
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