BME100 s2017:Group5 W8AM L4: Difference between revisions
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=OUR TEAM= | =OUR TEAM= | ||
{| style="wikitable" width="700px" | {| style="wikitable" width="700px" height="700px" | ||
|- valign="top" | |- valign="top" | ||
| [[Image: | | [[Image:Anisa_Ahamed.jpg|100px|thumb|Name: Anisa Ahamed]] | ||
| [[Image: | | [[Image:Jenna_Forrey.jpg|100px|thumb|Name: Jenna Forrey]] | ||
| [[Image: | | [[Image:Smita_Gopalakrishnan.jpg|100px|thumb|Name: Smita Gopalakrishnan]] | ||
| [[Image: | | [[Image:Matthew_Gruzda.jpg|100px|thumb|Name: Matthew Grudza]] | ||
| [[Image: | | [[Image:Fernanda_Nunez.jpg|100px|thumb|Name: Fernanda Nunez]] | ||
| [[Image:Luis_Paez.jpg|100px|thumb|Name: Luis Paez | | [[Image:Luis_Paez.jpg|100px|thumb|Name: Luis Paez]] | ||
|} | |} | ||
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* Lab coat and disposable gloves | * Lab coat and disposable gloves | ||
* PCR reaction mix, 8 tubes, 50 μL each: Mix contains Taq DNA polymerase, MgCl 2, and dNTP’s | * PCR reaction mix, 8 tubes, 50 μL each: Mix contains Taq DNA polymerase, MgCl 2, and dNTP’s | ||
* DNA/ primer mix, 8 tubes, 50 μL each: Each mix contains a different template DNA. All tubes | * DNA/ primer mix, 8 tubes, 50 μL each: Each mix contains a different template DNA. All tubes have the same forward primer and reverse primer | ||
have the same forward primer and reverse primer | |||
* A strip of empty PCR tubes | * A strip of empty PCR tubes | ||
* Disposable pipette tips: only use each only once | * Disposable pipette tips: only use each only once. Never reuse disposable pipette tips. If you do, the samples will become cross-contaminated | ||
* Cup for discarded tips | * Cup for discarded tips | ||
* Micropipettor | * Micropipettor | ||
* OpenPCR machine | * OpenPCR machine: shared by two groups | ||
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'''PCR Reaction Sample List'''<br> | '''PCR Reaction Sample List'''<br> | ||
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'''DNA Sample Set-up Procedure''' | '''DNA Sample Set-up Procedure''' | ||
<!-- In your own words, write an easy-to-comprehend list of the steps your group took to set up the PCR reaction in the PCR reaction tubes. End with placing the tubes in to the thermal cycler. Do not copy-paste the instructions from the Workbook. That will be considered plagiarism. --> | <!-- In your own words, write an easy-to-comprehend list of the steps your group took to set up the PCR reaction in the PCR reaction tubes. End with placing the tubes in to the thermal cycler. Do not copy-paste the instructions from the Workbook. That will be considered plagiarism. --> | ||
# | #Label each of the 8 tubes with the names listed above | ||
# | #Pipette 50 µL of the PCR reaction mix into the positive control tube | ||
# | #Dispose of the pipette tip to avoid cross-contamination | ||
#Pipette 50 µL of the DNA/primer mix into the positive control tube (total volume in tube is now 100 µL) | |||
#Repeat steps 2-4 for the rest of the tubes with designated DNA/primer mix for each | |||
#Close the lids on tubes and insert them into thermal cycler | |||
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'''Background: About the Disease SNP''' | '''Background: About the Disease SNP''' | ||
<!-- INSTRUCTIONS: This content is from PCR Lab B. Write a summary, at least five sentences long, about the disease SNP in your own words. --> | <!-- INSTRUCTIONS: This content is from PCR Lab B. Write a summary, at least five sentences long, about the disease SNP in your own words. --> | ||
SNP, which stands for “single nucleotide polymorphism”, is a disease that causes a genetic alteration common in many individuals. The disease occurs when there is a single change in the nucleotide, or a building block of DNA consisting of Adenine, Thymine, Guanine, and Cytosine. On average, SNP is located around every 300 nucleotides, which is a total of 10 million in the human genome. In our lab, we observed a variation of SNP located on the 7:117587799 chromosome. This specific SNP was linked to cystic fibrosis. | |||
"What Are Single Nucleotide Polymorphisms?" ''U.S. National Library of Medicine.'' National Institutes of Health, 21 Mar. 2017. Web. 21 Mar. 2017. | |||
'''Primer Design and Testing''' | '''Primer Design and Testing''' | ||
<!-- INSTRUCTIONS: Write a short summary of the results of your primer test. Underneath your summary, include a screen capture of the results web page. You may crop the image so that it only includes the relevant information. --> | <!-- INSTRUCTIONS: Write a short summary of the results of your primer test. Underneath your summary, include a screen capture of the results web page. You may crop the image so that it only includes the relevant information. --> | ||
For this lab, two sets of primers were designed, the non-disease primers and the disease-specific primers. The non-disease forward primer was written from the position of the SNP for a sequence of 20 bases long, and its reverse primary was written till the base that was 200 base pairs to the right of the SNP. Then, the disease primers were written by changing the final base of the non-disease forward primer to match the disease SNP nucleotide. | |||
We ran two different tests with the non-disease and the disease-specific primers. From the tests, we observed that the non-disease primers resulted in a match with the same chromosome as the chromosome we found earlier, confirming our results. In addition, the disease primers produced no matches, since the diseased codon would mutate at the AGT sites. | |||
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[[Image: BME100_Fig4.2.jpg]] | |||
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[[Image: BME100_Lab4.3.jpg]] | |||
Latest revision as of 22:06, 21 March 2017
BME 100 Spring 2017 | Home People Lab Write-Up 1 | Lab Write-Up 2 | Lab Write-Up 3 Lab Write-Up 4 | Lab Write-Up 5 | Lab Write-Up 6 Course Logistics For Instructors Photos Wiki Editing Help | |||||||||||||||||||||||||||||||||
OUR TEAMLAB 4 WRITE-UPProtocolMaterials
PCR Reaction Sample List
OpenPCR program
Research and DevelopmentPCR - The Underlying Technology Components on a PCR Reaction
Components during each step of the thermal cycling
Template DNA: Separates into two strands.
Template DNA: The two single strands tend to go back together but the abundance of primers in the tube prevents them of doing this.
Template DNA: DNA is now bind by the primers and Taq Polymerase, ready to be synthesized.
Template DNA: Each single-strand is synthesized in a copy of the original double stranded DNA molecule.
Template DNA: There is now two DNA molecules for each original template DNA at each PCR cycle, and the synthesized molecules are stabilized at the low temperature. Nucleotide Base Pairing
Base Pair Occurrence
SNP Information & Primer DesignBackground: About the Disease SNP SNP, which stands for “single nucleotide polymorphism”, is a disease that causes a genetic alteration common in many individuals. The disease occurs when there is a single change in the nucleotide, or a building block of DNA consisting of Adenine, Thymine, Guanine, and Cytosine. On average, SNP is located around every 300 nucleotides, which is a total of 10 million in the human genome. In our lab, we observed a variation of SNP located on the 7:117587799 chromosome. This specific SNP was linked to cystic fibrosis. "What Are Single Nucleotide Polymorphisms?" U.S. National Library of Medicine. National Institutes of Health, 21 Mar. 2017. Web. 21 Mar. 2017.
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