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Team:Calgary/11 June 2009
From 2009.igem.org
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| - | + | Gradient PCR Take Two | |
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| - | + | *Objective: To biobrick LuxOD47E in the 2.1pCR-TOPO vector. | |
| + | *Diluted LuxOD47E template to 40 ng/uL, (2.1uL template + 12.9 uL ddH20) | ||
| + | *Followed protocol from June 6th 2009 | ||
| + | *PCR Prodcuts visualized on a 1% agarose gel run at 120 V. See gel photo below. Lanes 1-12 contain mastermix + LuxOD47E in pCR2.1-TOPO vector template. Lane 13 is a negative control with mastermix + ddH2O. | ||
| + | [[Image:2009.06.12.LuxOD47E_Gradient_PCR.tif]] | ||
| + | *Analysis: There is considerable contamination in the negative control, however the gradient looks as expected. The source of contamination may possibly be from the primers that were shared between Viki and I. We will proceed to PCR purification anyways. | ||
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Revision as of 05:05, 21 August 2009
UNIVERSITY OF CALGARY
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CAROL
Descriptive Title of What You’re Doing
WIKI CODING HERE
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CHINMOYEE
CLASS
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EMILY
Gradient PCR Take Two
File:2009.06.12.LuxOD47E Gradient PCR.tif
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FAHD
Descriptive Title of What You're Doing
WIKI CODING HERE
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IMAN
Descriptive Title of What You're Doing
WIKI CODING HERE
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JAMIE
Descriptive Title of What You're Doing
WIKI CODING HERE
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JEREMY
Descriptive Title of What You're Doing
WIKI CODING HERE
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KATIE
Success
I was able to combine all three scripts into one by using the listen event and now it is possible for me to add more types of DNA to PCR and I can continue to easily add quite a few of them. I also was able to fix the error I was getting, which happened because as long as the first item stored in a list was found in the machine’s inventory, it believed everything was there, but when it went to do something with it, the item did not exist and an error occurred.
However, I would still like to change the way it is being used at this time since it is not really efficient to re-declare a variable every time you go through a loop. I will work on changing this at a later date.
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KEVIN
Plasmid Isolation of LuxCDABE (luciferase)
The LuxCDABE is being isolated by Caron Chan, but she needed some hands on this, and I was willing to help. LuxCDABE was isolated with Quigen's EndoFree Plasmid Maxi (quigen) according to the specification of the provider. A different kit than the one used in previous experiments was used because the LuxCDABE is very big in size (6kb), and the Minprep would not be capable of isolating the plasmid at a high enough concentration.
Despite of the use of Maxiprep, however, the spectrophotometer seemed to have not picked up any plasmid in the isolated sample, as the purity and concentration were below acceptable. Another Maxiprep may be needed to be done. |
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MANDY
Descriptive Title of What You're Doing
WIKI CODING HERE
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PATRICK
Descriptive Title of What You're Doing
WIKI CODING HERE
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PRIMA
Descriptive Title of What You're Doing
WIKI CODING HERE
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STEFAN
Descriptive Title of What You're Doing
WIKI CODING HERE
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VICKI
Re-do of gradient PCR because of contamination issues
Purpose: The last gradient PCR for BioBrick amplification failed because of contamination. This is the second attempt.
Materials and methods: Please refer to the June 4th entry for the observed protocol. Results: Shown below, with the lane key included. The blank negative control lane is reassuring. The ladder is a GeneElute 1kb Plus DNA ladder.
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