Editing
Alice:Single/2015-4-30
(section)
Jump to navigation
Jump to search
Warning:
You are not logged in. Your IP address will be publicly visible if you make any edits. If you
log in
or
create an account
, your edits will be attributed to your username, along with other benefits.
Anti-spam check. Do
not
fill this in!
===cDNA amplification by LD PCR=== :- Thaw all the reagents needed for PCR (except the enzyme) on ice. Gently vortex each reagent tube to mix and spin down briefly. Store on ice. :- Prepare enough PCR Master Mix for all the reactions, plus 10% of the total reaction mix volume. Combine the following reagents in the order shown. :- NOTE: Remove the DNA polymerase from the freezer, gently mix the tube without vortexing, and add to the Master Mix just before use. Mix the Master Mix well by vortexing gently and spin the tube briefly in a microcentrifuge to collect the contents at the bottom of the tube: :{| {{table}} border = 1 | align="center" style="background:#f0f0f0;"|'''volume per reaction''' | align="center" style="background:#f0f0f0;"|'''volume for 3 reactions''' | align="center" style="background:#f0f0f0;"|'''component''' |- | 12.5 ul|| 37.5 ul|| 2X SeqAmp PCR Buffer |- | 0.5 ul|| 1.5 ul|| PCR Primer II A - v3 (12 μM) |- | 0.5 ul|| 1.5 ul|| Nuclease-free water |- | 1.5 ul|| 4.5 ul|| SeqAmp DNA Polymerase |- | 15 ul|| 45 ul || Total Volume added per reaction |} :- Mix well and briefly spin in a minicentrifuge to collect the contents at the bottom of the tube(s). :- Place the tube(s) in a preheated thermal cycler with a heated lid and run the following program: :- 95°C 1 min -> ( 98°C for 10 sec -> 65°C for 30 sec -> 68°C for 3 min ) x 40 cycles -> 72°C 10 min
Summary:
Please note that all contributions to ZhangLabWiki may be edited, altered, or removed by other contributors. If you do not want your writing to be edited mercilessly, then do not submit it here.
You are also promising us that you wrote this yourself, or copied it from a public domain or similar free resource (see
ZhangLabWiki:Copyrights
for details).
Do not submit copyrighted work without permission!
Cancel
Editing help
(opens in new window)
Navigation menu
Personal tools
Not logged in
Talk
Contributions
Create account
Log in
Namespaces
Page
Discussion
English
Views
Read
Edit
View history
More
Search
Navigation
Main Page
Current events
Recent changes
Random page
Investigators
Matt Cai
Song Chen
Eric Chu
Dinh Diep
Elizabeth Duong
Shicheng Guo
Alan Fung
Daniel Jacobsen
Blue Lake
Huy Lam
Alice Li
Andrew Richards
Brandon Sos
Chris Wei
Yan Wu
Kun Zhang
Tools
What links here
Related changes
Special pages
Page information