Difference between revisions of "840:153g:Projects/project23/2012/10/25"

From OpenWetWare
(Continuation of the Mutagenesis Reaction)
(Continuation of the Mutagenesis Reaction)
Line 8: Line 8:
 
==Continuation of the Mutagenesis Reaction ==
 
==Continuation of the Mutagenesis Reaction ==
  
* On Tuesday, we ran the gel for last week's PCR products.  This was done on a dual-lane chamber, loaded as follows:
+
* On Tuesday, we ran the gel for last week's PCR products.  The high concentration of 1:20 template DNA, and a low concentration of 1:200 were prepared with water for each the upstream and downstream fragments.  Our nomenclature used is three-part, and outlined below.
  
 +
*1.)Primer configuration
 +
**N = no primers used
 +
**P = outside primers used
 +
*2.)Template DNA concentration
 +
** + = 1:20
 +
** - = 1:200
 +
*3.)Annealing temperature for PCR, in °C
 +
 +
 +
*Our positive control was standard template DNA and associated primers from lab stock
 +
*Our negative control had outside primers and no template
 +
 +
 +
*We loaded our samples as follows:
  
 
*Top Lane:
 
*Top Lane:
Line 28: Line 42:
 
#N+60
 
#N+60
 
#N+65
 
#N+65
#Positive Control
+
#Negative Control
 
#N-55
 
#N-55
 
#N-60
 
#N-60

Revision as of 15:14, 29 October 2012

Owwnotebook icon.png Cloning of the OmcF gene <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>

Continuation of the Mutagenesis Reaction

  • On Tuesday, we ran the gel for last week's PCR products. The high concentration of 1:20 template DNA, and a low concentration of 1:200 were prepared with water for each the upstream and downstream fragments. Our nomenclature used is three-part, and outlined below.
  • 1.)Primer configuration
    • N = no primers used
    • P = outside primers used
  • 2.)Template DNA concentration
    • + = 1:20
    • - = 1:200
  • 3.)Annealing temperature for PCR, in °C


  • Our positive control was standard template DNA and associated primers from lab stock
  • Our negative control had outside primers and no template


  • We loaded our samples as follows:
  • Top Lane:
  1. 100 bp ladder
  2. P+55
  3. P+60
  4. P+65
  5. Positive Control
  6. P-55
  7. P-60
  8. P-65
  9. 100 bp ladder


  • Bottom Lane:
  1. 100 bp ladder
  2. N+55
  3. N+60
  4. N+65
  5. Negative Control
  6. N-55
  7. N-60
  8. N-65
  9. 100 bp ladder


  • We ran these on a 1% agarose gel for 30min at 120V


  • Results: