Madhadron:Cruft: Difference between revisions

Revision as of 14:32, 7 February 2007

My own personal brand of labelling obsessiveness:

DNA shall hav a light green dot on the top of it's tube if it is a maxiprep; bright orange if it is primer; and I haven't had to assign a miniprep color yet.
Chemicals will be labelled on orange tape if sterile, and on green if they are not.
E. coli stock will have a dark green dot on the top of its tube. M. smegmatis will have dark blue.

Neeraj's advice for revising my PCR:

Titrating Mg, DNA, etc. is for when you don't get any bands; no good if you have too many bands.
Use 30s instead of 1 minute for the three stages of PCR in general. Strongly binding primers will bind just fine in 30s, extension for 30s in 1 minute will reduce nonspecific large stuff, and 30s at denaturation will destroy less of the polymerase at each cycle.
Can try adding 5M DMSO if there's still a lot of nonspecific junk.

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 My own personal brand of labelling obsessiveness:
-* DNA shall half a light green dot on the top of it's tube if it is a maxiprep; bright orange if it is primer; and I haven't had to assign a miniprep color yet.
+* DNA shall hav a light green dot on the top of it's tube if it is a maxiprep; bright orange if it is primer; and I haven't had to assign a miniprep color yet.
 * Chemicals will be labelled on orange tape if sterile, and on green if they are not.
 * E. coli stock will have a dark green dot on the top of its tube.  M. smegmatis will have dark blue.
+Neeraj's advice for revising my PCR:
+* Titrating Mg, DNA, etc. is for when you don't get any bands; no good if you have too many bands.
+* Use 30s instead of 1 minute for the three stages of PCR in general.  Strongly binding primers will bind just fine in 30s, extension for 30s in 1 minute will reduce nonspecific large stuff, and 30s at denaturation will destroy less of the polymerase at each cycle.
+* Can try adding 5M DMSO if there's still a lot of nonspecific junk.