Editing
Jeff:LabNotes/Microbiome/2011-9-21
(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!
== '''Single Mammalian Cell Amplification: Ideas''' == *I want to apply my device to mammalian cells as well *It would be cool if we could use it for single stem cells or cancer cells '''Contamination''' *Since currently I have ~ 50% human contamination rate, I cannot trust the results from human cells *I will use mouse embryonic fibroblasts (MEFs) from the Xu lab *Alan will help me grow the first batch and hopefully I can take over maintaining this cell line '''Visualization''' *These cells are much bigger than bacterial cells *I can spread them, stain with SYBR green, and visualize to get a good idea of the proper concentration to use. '''Process''' *Everything can be done using the exact same method as with bacterial cells *The cell lysis will be different since there is no cell wall in mammalian cells. Freeze/thaw lysis will not be used *It would be advantageous to use a strong base/alkaline solution for cell lysis as this is needed for denaturing and will remove the need for adding additional reagents. *Based on previous papers, I can use something similar to an ALS buffer: 200 mM KOH, 50 mM DTT, and incubate at 65 C for 10 min *ALS mix: 4 uL 5M KOH, 5 uL 1M DTT, 91 uL water *Then use the following NS buffer: 900 mM Tris-HCl, 300 mM KCl, 800 mM HCl *NS mix: 4.5 uL 2 M Tris-HCl, 750 uL 4 M KCl, 66.67 uL 12N HCl, 4.683 mL water *The MDA, library construction process should be the same. '''Applications''' *Look at CNV's *Look at cell diversity at different embryonic stages
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