Editing
Dinh:Probes Prep
(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!
== PAGE size selection == * UV all gel running buffer and chamber. * UV shearing apparatus. ** Prepare 3x (2.0 mL tube + 0.5 mL tube) per gel. Cut off the caps of 2.0 mL tube to enable centrifuge at 15,000 rpm. * Puncture 0.5 mL gel with 20 gauge needle. * Run 6% gel for 40 minutes at 200 V. (Run lower volt to get better band resolution). * Add 1.5 mL TE buffer to sheared gel. * Rotate overnight at room temperature. * 50C for 15 minutes at 900 rpm on thermomixer. (Or 37C for 1 hour at 900 rpm). Yield with 37C/1 hour seems decent, not sure which is better. * Quickly spin-down tubes & filter through 1-2 Nanosep columns. Keep tube warm. * Pool flow-through in 1x 15 mL tube. * Add 2.5x 100% ethanol, 0.1x 3M sodium acetate, and 8 uL of glycoblue * Store in -80C for at least 1 hour. * Spin at >4000 rpm for 40 minutes at 4C * Remove supernatant & add 1 mL 80% ice-cold ethanol. Transfer to 1x 1.5 mL tube. * Spin at >10,000 rpm for 15 minutes at 4C * Remove supernatant & vacumm dry for 10-30 minutes * Add 30 uL of pre-warmed nuclease-free water. * QUBIT ssDNA to quantify yield.
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