cazmantis

That's great! Thanks everyone for the replies - eBay strikes again

Hi, I am not sure if this is in the right place or not! I need to obtain some 80% ethanol in order to preserve Hymenoptera specimens but I am having real trouble pinning down how to do this! I know you can procure absinthe up to 80% but I read that specimens should ideally be preserved in a clear alcoholic liquid. Could somebody please help me out with this query! I just need to make sure the DNA will be preserved for (a maximum of) 2 months! Thanks! Caz

Hi, Well things are going well with my project and I think I'm getting the hang of NCBI. However there is one little concern which is still niggling at me. Firstly I should explain that I am blasting genomic DNA from Nasonia vitripennis and Apis mellifera in order to find similarities between two gene sequences. However, when I look at the genomic DNA of one of the genes (NM_001013359) in the gene record it describes it as "complement", whilst the other sequence (NM_001161675) does not have the word "complement" next to the genomic DNA record. So here is my question - can I enter the FASTA sequences of these two stretches of genomic DNA into the "align 2 or more sequences" function of BLAST as is or must I translate one of them? (I'm not sure what "complement" menas but I was under the impression that it was on the negative strand and so it stands to reason that even though I am comparing 2 similar genes here they will be from opposite strands and so therefore BLAST may not find any similarities!) Screenshot attached. Any help would be most appreciated. I hope this was clear! Thanks, Caz NCBI issue screen.doc

That's great - thanks. Good to have a confirmation of what I had presumed! I am just trying to ascertain that the gene which codes for a particular protein (in this case I'm looking at antigen 5) is a product of the same locus in two species. It has been suggested that looking for similarities in the non-coding regions will help identify that. I am using Clustal also to align these sequences but it is hard for me to tell (due to lack of experience) exactly how similar means "similar enough"! Thanks for your help! Caz

Hi, Could I just check with someone if I have got this right please? My definition of a flanking region of a gene is the region x bp upstream from the start codon and the equivalent downstream from the stop codon? For the current gene I am looking at I am using about 500bp as my flanking region as it seemed a sensible figure (1,000 bp upstream is the end of another gene so I didn't want to go into that) but is there an official way to identify a flanking region or would my estimate be appropriate? I am BLASTING these regions to try and identify similarity. Thanks so much! Caz

Hi! Thanks for the replies, that's just great. Shortly after I posted I managed to figure out how to use the sequence viewer in NCBI a bit better and managed to export my gene plus the flanking regions for analysis in Clustal - but it's great to have the confirmation that I'm doing it right I think I have made way too much allowance for my flanking regions though - as per your suggestion I will cut it down to a few hundred base pairs. I'll have a look at the software which predicts placement of promotors too - if nothing else it will be interestingt o see what results I obtain. Thanks so much! Until next time Caz

Hi Charon, thanks so much for your swift reply. I have just a question or two from what you have said below (many apologies - I'm new at this and getting myself into a real muddle!) Finding the position in the genome of the gene I am looking at is proving difficult. I am using mainly Apis and Nasonia sequences so I have full genomes for them but I'm not sure how one quantifies "the position in the genome". Are we just talking about which chromosone and locus the gene is located on or should I try and be a little more precise? I think I have been working on the lines of what you suggest below. I do have some confusion about which part of the gene constitutes the "flanking region" and also how I extract the upstream and downstream regions in NCBI database? I will continue my reading but any help would be greatly appreciated! Best, Caz

Hiya, I am currently BLASTING genomic data in FASTA format between species to try and identify a conserved locus. I have obtained quite a good list of BLAST results, but what I am specifically looking for are matches in the flanking regions of the gene to make sure they are a product of the same locus. I have been looking into this for a week or so and my progress is...slow to say the least! I can blast the gene which specific mRNA of interest is derived from (I am using genomic DNA), and of course once I obtain a result I can see where the matches are in the nucleotide sequences. I am just really stuck at identifying where in the piece of genomic DNA I'm looking at the flanking region is! I am aware that it will be at the 5' end and will contain elements such as signal peptide and TATA box, but my lack of experience is really holding me back on figuring this out. Could anyone help me out with any pointers? Thanks so much! Caz

That's great - thanks so much for your help. Making a lot more sense now. Best, Caroline

Hiya! Wow thanks so much for the replies - very useful stuff so far and has served to illuminate my own lack of knowledge of the subject! I think I am getting a little confused with these things. The PREDICTED sequences I am looking at are nucleotide sequences and I am not sure how I would cross reference that against a genome. It just doesn't seem to make sense to me so I assume that my lack of experience in the field means I don't have access to all the facts! For example NCBI accession number XM_001120951 - it says this nucleotide sequence has been predicted from the genomic sequence. I am finding this quite confusing as surely the nucleotide exists or it doesn't. I want to understand how these predicted sequences are different to (let's say) "normal" sequences. I understand this may be a little in depth to expect an answer on but if anyone could perhaps reccomend a book which may cover this aspect of genomes that would be just as helpful for me. Thanks so much for your help, Caroline

Bit new to bioinformatics and just wondered how useful PREDICTED amino acid sequences derived from the genome are (I have found several in NCBI). If trying to BLAST these sequences to find conserved sequences, are the results going to be of use or is it better to not bother with these sequences at all? Thanks, Caz