Editing
Daniel:Notebook/HiResChrPaint/2013-7-3
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!
=S2 Probe Prep ([[Daniel:Notebook/HiResChrPaint/2013-5-29|Started 5/29/2013]])= [[Daniel:Notebook/HiResChrPaint|Back to Calendar]] ==ULYSIS Dye Coupling== Trying the ULYSIS dye labeling technique using [[Daniel:Notebook/HiResChrPaint/2013-6-10#Nanodrop Results|V4S2 sample without aadUTP]] as a trial run. Sample has been dried in a vacuum centrifuge and will need to be resuspended. '''V4S2 (no aa-dUTP) ~1.5 ug''' *Following [[Daniel:Protocols/AlexaDyeCouple#ULYSIS|ULYSIS protocol]] 1. Add 5 uL DMSO to Alexa dye 488 2. Resuspend DNA in 20 uL labeling buffer (Component C) 3. Denature DNA for 5 minutes at 95C; snap cool on ice 4. Add 1 uL dye to sample; add 4 uL labeling buffer 5. Incubate for 15 minutes at 80C, snap cool on ice 6. Purify using [[Daniel:Protocols/CentriSep|Centri-Sep Column]] ===Results=== {| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> |- style="font-size:12pt;font-weight:bold" align="center" | width="65" height="30" | ng/uL DNA | width="65" | ug DNA in 20 uL | width="65" | pmol/uL DNA | width="65" | pmol/uL dye | width="65" | dye/probe | width="65" | dye/100bp |- style="font-size:12pt" valign="bottom" | align="center" height="15" | 111.9 | align="center" | 2.2 | align="center" | 4.2 | align="center" | 1.9 | align="center" | 0.5 | align="center" | 0.62 |} Seems like this kit doesn't work as well as the ARES. The instructions say labeling efficiency should be around 44 fluorophores/67 bases (guanine only, actual labeling efficiency the way I calculate it should be around 16.4% assuming equal base probability). I'm going to look into possible reasons why this ratio is so low for me.
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