"
Team:Arizona State/Notebook/PCRLog
From 2011.igem.org
(Created page with "{{:Team:Arizona State/Templates/main|title=PCR Log for E. Coli Cas Genes|content= __NOTOC__ ==Overview== <p>This logbook is a record of the majority of our attempts to PCR ampli...") |
|||
| Line 7: | Line 7: | ||
===Notes=== | ===Notes=== | ||
<p>Suggested annealing temperatures are based on [http://www.neb.com/nebecomm/tech_reference/TmCalc/Default.asp NEB Tm Calculator], as called for in the [http://www.neb.com/nebecomm/tech_reference/polymerases/phusion_high.asp?&utm_source=Google&utm_medium=CPC&utm_term=+phusion&utm_campaign=Phusion NEB Phusion DNA Polymerase] protocol.</p> | <p>Suggested annealing temperatures are based on [http://www.neb.com/nebecomm/tech_reference/TmCalc/Default.asp NEB Tm Calculator], as called for in the [http://www.neb.com/nebecomm/tech_reference/polymerases/phusion_high.asp?&utm_source=Google&utm_medium=CPC&utm_term=+phusion&utm_campaign=Phusion NEB Phusion DNA Polymerase] protocol.</p> | ||
| + | <p>Desired band for CasABCDE at ~4300bp</p> | ||
| + | <p>Desired band for Cas3 at ~2667bp</p> | ||
==Primer Round 1== | ==Primer Round 1== | ||
| Line 14: | Line 16: | ||
<p>Gel Results:</p> | <p>Gel Results:</p> | ||
[[Image:]] | [[Image:]] | ||
| - | + | CasABCDE: Very faint bands near target length | |
| + | Cas3: No bands | ||
==Primer Round 2== | ==Primer Round 2== | ||
Revision as of 21:31, 28 September 2011
|
|
 OverviewThis logbook is a record of the majority of our attempts to PCR amplify CRISPR-associated (Cas) ABCDE and Cas 3 of E. coli K12 MG1655. The record begins after our attempt to individually PCR amplify each of the 6 genes, which was unsuccessful. Here, three different sets of primers were used to attempt PCR amplification of the Cas genes in two sections. NotesSuggested annealing temperatures are based on [http://www.neb.com/nebecomm/tech_reference/TmCalc/Default.asp NEB Tm Calculator], as called for in the [http://www.neb.com/nebecomm/tech_reference/polymerases/phusion_high.asp?&utm_source=Google&utm_medium=CPC&utm_term=+phusion&utm_campaign=Phusion NEB Phusion DNA Polymerase] protocol. Desired band for CasABCDE at ~4300bp Desired band for Cas3 at ~2667bp Primer Round 1July 17, 2011CasABCDE: Touchdown PCR, Start Temp 70, -0.2 / cycle, Final Temp 64</p>
Gel Results: [[Image:]] CasABCDE: Very faint bands near target length Cas3: No bands Primer Round 2==Primer Round 3== |
