Muratore:Protocols/PCR/Basic

From OpenWetWare

(Difference between revisions)
Jump to: navigation, search
(New page: {|border="1" |+ Basic PCR protocol |- !Tube !!sterile H<sub>2</sub>O !!Pfu Buffer !!Template !!5'primer !!3'primer !!dNTPs !!Pfu Turbo !!wax |- ! !! !!(10X) !!(10 μg/mL) !!(4 μM) !!(4 μ...)
Line 1: Line 1:
{|border="1"
{|border="1"
|+
|+
-
Basic PCR protocol
+
'''Basic PCR protocol'''
|-
|-
!Tube !!sterile H<sub>2</sub>O !!Pfu Buffer !!Template !!5'primer !!3'primer !!dNTPs !!Pfu Turbo !!wax
!Tube !!sterile H<sub>2</sub>O !!Pfu Buffer !!Template !!5'primer !!3'primer !!dNTPs !!Pfu Turbo !!wax
Line 7: Line 7:
! !! !!(10X) !!(10 μg/mL) !!(4 μM) !!(4 μM) !!(10 mM ea) !!(2.5 U/μL) !!
! !! !!(10X) !!(10 μg/mL) !!(4 μM) !!(4 μM) !!(10 mM ea) !!(2.5 U/μL) !!
|-
|-
-
!A
+
!Experimental
|72 μL ||10 μL ||5 μL ||5 μL ||5 μL ||2 μL ||1 μL ||50 μL
|72 μL ||10 μL ||5 μL ||5 μL ||5 μL ||2 μL ||1 μL ||50 μL
|-
|-
-
!B
+
!(-) Control
|73 μL ||10 μL ||5 μL ||5 μL ||5 μL ||2 μL ||0 μL ||50 μL
|73 μL ||10 μL ||5 μL ||5 μL ||5 μL ||2 μL ||0 μL ||50 μL
|}
|}

Revision as of 13:42, 26 May 2011

Basic PCR protocol
Tube sterile H2O Pfu Buffer Template 5'primer 3'primer dNTPs Pfu Turbo wax
(10X) (10 μg/mL) (4 μM) (4 μM) (10 mM ea) (2.5 U/μL)
Experimental 72 μL 10 μL 5 μL 5 μL 5 μL 2 μL 1 μL 50 μL
(-) Control 73 μL 10 μL 5 μL 5 μL 5 μL 2 μL 0 μL 50 μL
  • Add each from left to right. Mix and spin before adding Pfu Turbo. Gentle mixing and spin after Pfu Turbo. Don't mix in wax.
  1. 5' @ 95°C
  2. 30" @ 95°C
  3. 2' @ Tm-5°C
  4. 2' @ 78°C
  5. repeat steps 2-4 x 29 more times
  6. 5' @ 78°C
Personal tools