Search found 12 matches

by dustman
Mon Jun 25, 2012 2:09 pm
Forum: Molecular Biology
Topic: PCR from cDNA question
Replies: 1
Views: 6129

Disclaimer: I normally work with plasmid/genomic DNA, so might be wrong in my suggestions. Possible problems may be: 1. low amount of mRNA. You don't have that many cells total and you can't be sure that mRNA is stable for your genes of interest. Possible solution -- increase number of cycles. 2. Pr...
by dustman
Tue Jun 05, 2012 8:05 am
Forum: Molecular Biology
Topic: RFLP Question
Replies: 1
Views: 5932

According to your gels, which are bit poorly described in the post, Bob is AS ans Sally AA. If both genes, M and D are located in the same chromosome, chances are ~25/50% that a kid gets AS and 0% to get anemia, unless one gene mutation is enough for its development. This is based on Mendel Btw, wit...
by dustman
Sat May 26, 2012 11:04 pm
Forum: Molecular Biology
Topic: Do the simplest bacteria have ribosomes and helicase?
Replies: 27
Views: 25367

Daemach, you must check chemical reactions in RNA synthesis before asking any more questions. Here's a link with a pic of actual nucleotide incorporation into oligo chain: m Thing is, A, C, G and T/U in DNA/RNA molecule mean separate nuclotides, with purine/pyrimidine, carbohydrate and exactly one p...
by dustman
Sat May 26, 2012 7:08 am
Forum: Molecular Biology
Topic: Do the simplest bacteria have ribosomes and helicase?
Replies: 27
Views: 25367

Re:

1) I'm not sure what this means. 2) Do you have any links? 3) DNA does have to be strung together in a perfect sequence to create a cell that functions correctly. And my textbook says that had to happen for millions of generations for evolution to work. Am I missing something? 1. Not to be rude but...
by dustman
Fri May 25, 2012 2:54 pm
Forum: Molecular Biology
Topic: restriction reaction on agarose gel slices
Replies: 2
Views: 2647

restriction reaction on agarose gel slices

I'm thinking of reducing steps for insert production. ATM I have following steps: 1. PCR 2. agarose gel with PCR products 3. gel extraction of insert of interest 4. restriction reaction 5. enzyme removal <- I often use KpnI/BamHI and can't go with heat inactivation 6. ligation with backbone and tran...
by dustman
Fri May 25, 2012 2:13 pm
Forum: Molecular Biology
Topic: Do the simplest bacteria have ribosomes and helicase?
Replies: 27
Views: 25367

You want a simple answer to a very hard question that hasn't being properly answered in decades? If you want to speculate, just think of a liposome (bilayer of lipids), with some RNA and nucleotides inside. Just for your info, ATP is RNA. Give it time (billions of years), numbers (huge), additional ...
by dustman
Thu May 24, 2012 10:57 am
Forum: Molecular Biology
Topic: what is DNA & RNA?
Replies: 9
Views: 8766

Re:

vinayaksabnis wrote:Both are nucleic acids containing genetic information with different sugars deoxyribose and ribose sugars



RNA is more than 'genetic information'. tRNA, microRNA, ribosomes and so on have plethora of other functions. Only mRNA and genome of some viruses falls in 'genetic information' type.
by dustman
Thu May 24, 2012 10:47 am
Forum: Molecular Biology
Topic: ligation problem although try everything,suggestions?
Replies: 7
Views: 7760

Btw, read transformation protocol carefully. You incubate specifically treated bacs with your ligation reaction and heat shock exactly as stated in the protocol by Invitrogen. 30 s at 42C is not taken out of blue. SOC is used for cells to recover before they are exposed to antibiotics in agar. You c...
by dustman
Thu May 24, 2012 10:42 am
Forum: Molecular Biology
Topic: ligation problem although try everything,suggestions?
Replies: 7
Views: 7760

You don't say what you do after restriction. Do you remove restriction enzymes? Or simply kill restrictazes by heat-shocking? Do you treat digested vector with CIAP (phosphotase)? If you simply heat-shock restriction reaction and proceed with ligation, it can cause your problem. Several enzymes, lik...
by dustman
Thu May 10, 2012 10:41 am
Forum: Molecular Biology
Topic: Integrated software tools
Replies: 4
Views: 4943

Re: Re:

What exactly is in silico PCR and virtual cloning? You pick template file, primer(s), specify conditions, run and get a fragment of certain length with expected nucleotide sequence, so you don't need to do it manually <- in silico PCR. Sometimes provides only fragment size(s). Pick two DNA sequence...
by dustman
Thu May 10, 2012 9:45 am
Forum: Molecular Biology
Topic: Integrated software tools
Replies: 4
Views: 4943

Thank you, Patrick,

Geneious looks interesting, at least after one day trial. Main thing I'm missing is automated plasmid annotations using existing database (or at least I haven't found this feature), a la PlasMapper. I love transparent UniProt/NCBI integration into software.
by dustman
Tue May 08, 2012 12:17 pm
Forum: Molecular Biology
Topic: Integrated software tools
Replies: 4
Views: 4943

Integrated software tools

I'm trying to find software package that can do all of the following things in transparent or semi-transparent way: Multiple alignment Restriction analysis Sequencing view Primer design in silico PCR Mutagenesis Virtual cloning Plasmid maps UGENE can do most of these things but its focus on 'project...