Search found 43 matches

by Ammie
Thu Aug 06, 2009 10:51 am
Forum: Molecular Biology
Topic: 3' RACE no polyA tail???
Replies: 0
Views: 1674

3' RACE no polyA tail???

Would be happy if someone could help me: I want to do a 3' RACE on a sequence without a poly A tail- how can I do it?
by Ammie
Tue May 20, 2008 6:53 am
Forum: Cell Biology
Topic: co-transfection- how does it work?
Replies: 0
Views: 7175

co-transfection- how does it work?

Earlier, I did a project as a part of my education where I did co-transfection with two plasmids, one containing GFP and one the gene of interest. The GFP was the transfection control and we only looked closer to the cells that were green and the theory was that the cells that took up the GFP-plasmi...
by Ammie
Fri Apr 18, 2008 1:52 pm
Forum: Molecular Biology
Topic: How would you create a strand specific PCR?
Replies: 0
Views: 2210

How would you create a strand specific PCR?

Hi! I want to run a strandspecific PCR. I have sense and antisense RNA transcript as control. How would you do it? The simpliest would be just to use one primer for cDNA synthesis (forward får antisense and reverse for sense) and than do a real time PCR with that primer pair. In theory that would wo...
by Ammie
Mon Mar 10, 2008 7:51 am
Forum: Molecular Biology
Topic: Got a gel extraction kit good for sequencing?
Replies: 3
Views: 1202

I used Promega gel and pcr-product clean up kit, and that have worked for me.
by Ammie
Mon Feb 18, 2008 3:11 pm
Forum: Molecular Biology
Topic: RNase protection assay and agarose gel
Replies: 2
Views: 2051

Thanks for your answer blcr11! I have made a first try now, with a RNA transcript insted of total RNAs from cells as a pre-trial. And it works at least with agarose gel! the PAGE will give a sharper band, but I think that it is sharp enough now. But how about the sensitivity? Could the detection lim...
by Ammie
Mon Feb 18, 2008 3:09 pm
Forum: Molecular Biology
Topic: How to know how to do the electroporation?
Replies: 1
Views: 1817

How to know how to do the electroporation?

Hi! Does anyone know how I can find out what parameters that is good to use for my cell line (PLC/PRF/5) Have tried to google it for a while but do not find anything. Thanks!
by Ammie
Thu Feb 14, 2008 2:30 pm
Forum: Molecular Biology
Topic: RNase protection assay and agarose gel
Replies: 2
Views: 2051

RNase protection assay and agarose gel

From Ambion technical bulletin #169: "Northern assays reqire the total RNA to be resolved on a denaturing agarose gel first, then transferred to a membrane and immobilized for subsequent hybridization. Nonisotopic RPAs which utilize probes labeled with modifyed nucleotides are transferred to a ...
by Ammie
Wed Dec 05, 2007 2:34 pm
Forum: Molecular Biology
Topic: About DNA & RNA
Replies: 2
Views: 3151

I would say 1H, 2G, 3E, 4C, 5A, 6D, 7B, 8F.
Like MrMistery
by Ammie
Fri Oct 05, 2007 12:06 pm
Forum: Molecular Biology
Topic: RNA storage
Replies: 3
Views: 1907

a follow-up question...
... how long at -20?
by Ammie
Wed Oct 03, 2007 11:08 am
Forum: Molecular Biology
Topic: Primers for QuikChange - desalted vs PAGE purified
Replies: 2
Views: 2678

Hi! I used desalted primers for multi QuikChange, and out of three sequenced colonies I got one with four of five mutations and the other with three of five. My primers were all 40nt long.

Maybe it would have worked even better with page-purifyed primers.
by Ammie
Tue Oct 02, 2007 3:05 pm
Forum: Molecular Biology
Topic: Best way to induce several mutations?
Replies: 2
Views: 1498

I had 5 that were spread over around 6000kb.

I decided to go with Multi QuikChange from Stratagene
by Ammie
Tue Oct 02, 2007 3:02 pm
Forum: Molecular Biology
Topic: QuikChange® Site-Directed Mutagenesis Kit
Replies: 3
Views: 3058

Not really a answer to your question but... I have used Multi QuikChange kit from Stratagene where you can mutate up to 5 nt's on the same time. When I sequenced 3 colonies, two of them had 3 of 5 mutations and one hade 4 of 5 mutations. I will now sequence 3 more colonies and hope to find one that ...