Editing
Capture Test of CES36k18bp
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!
===Capture Protocol=== '''Probe and Sample''' Probes: CES36k18bp, 5.6ng/ul Sample is Jurkat DNA standard, 100ng/ul (2.00ul) The Probe:Target ratio is 100:1 SLN Mix: Stoffel 2U/ul + AMPLigase 4U/ul + dNTP 0.25mM '''Calculations''' Probe: CES36k18bp: 5.6ng/ul; 36,000 probes; 170nM Volume of CES36k18bp needed = (200ng/3pg * 100 * 36000)/(6.0 * 10^23 * 170 nM) = 2.35ul '''Reaction Setup''' Jurkat DNA: 2.00ul 10X AMPLigase Buffer: 1.50ul CES36k18bp: 2.35ul H2O: 14.15ul 94C, 5min -> -0.1C/sec to 60C -> 60C, 24h -> add 2ul SLN mix -> 60C, 24h -> 95C, 2min -> 37C, 1min -> add 2ul Exo I&III -> 37C, 2h -> 94C, 5min -> 4C hold '''PCR''' Template: 8ul 2X Phusion mix: 50ul 100uM AmpF6.3NH2: 0.2ul 100uM AmpR6.3NH2: 0.2ul 50X SYBG: 0.4ul H2O: 30ul 98C, 30sec -> (98C, 10sec -> 58C, 20sec -> 72C, 20sec -> read) x 20 -> 72C, 3min -> 4C hold ==Results== The post-capture PCR was allowed to run to completion. The product from this PCR, 100ul total, was then run through a TBE gel to check for viable product. '''Gel Setup''' 25bp Ladder: 0.5ul Ladder + 3ul 1X TE Solution + 3ul 6X Loading Dye Low Mass Ladder: 0.5ul Ladder + 3ul 1X TE Solution + 3ul 6X Loading Dye Sample (x2): 5ul Sample (PCR Product) + 5ul 6X Loading Dye The TBE gel was run at 250 volts for 20 min: [[image: CES_Capture_Test_Annotated.jpg]]
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