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humalog's Content

There have been 22 items by humalog (Search limited from 14-April 20)


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#109120 How am I supposed to view/handle really long nt sequences on NCBI?

Posted by humalog on 07 May 2011 - 09:42 AM in Bioinformatics and Biostatistics

Thanks HomeBrew I am processing your reply now.



#108869 How am I supposed to view/handle really long nt sequences on NCBI?

Posted by humalog on 05 May 2011 - 02:48 AM in Bioinformatics and Biostatistics

Another thing, is it just me or has the NCBI Nucleotide turned VERY much slower than what it was about 1 year ago, in terms of retrieving nt sequences.
I've noticed this on two different otherwise-fine internet networks.

I don't even have the patience to wait for a sequence to download (it takes several minutes) while the same sequence would download in a few seconds some months ago.
:(



#108861 How can I find genomic sequence of a transcript

Posted by humalog on 05 May 2011 - 12:28 AM in Bioinformatics and Biostatistics

Hi
I haven't used bioinformatics for a bit so I need some memory refreshement.

I know how to find the nt sequence of a transcript. (on NCBI Nucleotide)
But how can I find the genomic sequence that codes for it (in other words the sequence on the genome that encodes it)?
Specifically I need this, so I can find the flanking sequences of a gene.



#108855 How am I supposed to view/handle really long nt sequences on NCBI?

Posted by humalog on 04 May 2011 - 11:32 PM in Bioinformatics and Biostatistics

Hi.
Say that I want to run a really long sequences through some bioinformatics software.
If I find this sequence on NCBI nucleotide, for example, and press on FASTA to view its sequence, the NCBI site takes ages to download/show the sequence, because its reallllllly long.
Also, even if it did download (something that didn't happen because I never had the patience for) I'm not sure whether I'd be able to copy such a long sequence on my clipboard, and then how would the other bioinfo software be able to process such a long sequence.
Is there an easier way to handle so large sequences?
I'm thinking whether I could download the FASTA sequence as a file instead? Would that make any difference? And how would I go about doing that?



#85478 Do the transcript nucleotide sequences in nucleotide databases include 3'UTR

Posted by humalog on 01 September 2010 - 11:42 AM in Bioinformatics and Biostatistics

Another related question I have is this:

Should I expect the end to a transcript sequence that I find in a database to contain the poly(A) signal somewhere near its end? Or could the poly(A) signal be more downstream from its end, and therefore not shown in this sequence?



#85476 Do the transcript nucleotide sequences in nucleotide databases include 3'UTR

Posted by humalog on 01 September 2010 - 11:29 AM in Bioinformatics and Biostatistics

But what about the cases where there seems to be no poly(A) tail at the end of a transcript?

For example http://www.ncbi.nlm....93?report=fasta

Also, in the example I gave, what is this long AAAAAAAAAAAA.... rich region in its beginning and the long TTTTTTTT... at its end?

thanks



#85469 How many nt apart are the promoter from the Start codon?

Posted by humalog on 01 September 2010 - 09:48 AM in Bioinformatics and Biostatistics

thanks



#85389 How many nt apart are the promoter from the Start codon?

Posted by humalog on 31 August 2010 - 08:11 PM in Bioinformatics and Biostatistics

Hi

I wanted to ask in regards to the promomer region and the start codon.

In what proximity are they with each other? How many nt close can I expect them to be?

thanks



#85213 How many bp downstream of Stop codon is the poly(A) signal?

Posted by humalog on 30 August 2010 - 10:38 AM in Bioinformatics and Biostatistics

Hi

In regards to stop codons and poly(A) signals, how many nucleotides downstream (>3') should the poly(A) signal be expected to be from the stop codon?

-Or should it be upstream?-

My poly(A) signal is TATAAA.

If you are lazy, appropriate reference to an informative website would also be appreciated

Thanks!.



#85048 If a 20bp primer differs from template DNA by 20bp, can PCR work?

Posted by humalog on 28 August 2010 - 01:26 PM in PCR, RT-PCR and Real-Time PCR

Hi

I was trying to PCR-amplify using a primer of around 20bp and it work.
Now when I check my data it seems to me that the primer they gave me to work with has a difference of 7bp in relation to the template DNA.

I wanted to ask if 7bp difference between primer and DNA template should normally make the PCR not work. In this case I might have entered the data (the primer sequence) in my notebook incorrecly.
Or would the PCR still work with the 7bp difference?



#84158 Repost: A nice Word Addon for sequence manipulation and analysis

Posted by humalog on 19 August 2010 - 04:39 PM in Bioinformatics and Biostatistics

Wow this is unbelievably helpful!



#84154 Do the transcript nucleotide sequences in nucleotide databases include 3'UTR

Posted by humalog on 19 August 2010 - 03:31 PM in Bioinformatics and Biostatistics

Yes, 3'UTR is included in mRNA sequence or CDs. In your example, you can find the polyA tail in the sequence.



Yes. Thanks.



#84125 Do the transcript nucleotide sequences in nucleotide databases include 3'UTR

Posted by humalog on 19 August 2010 - 11:12 AM in Bioinformatics and Biostatistics

Hi

I wanted to ask:

When we find a transcript sequence, for example in NCBI nucleotide database, is the 3' UTR regions supposed to be included in this sequence?

Let's take for example the mouse p53 gene variant 2, the one on this link: http://www.ncbi.nlm..../NM_001127233.1). Let's look at it's nucleotide sequence: Does it include the 3' UTR region (i.e. the polyadenylation signal, the stop codon, etc) ?

Please explain me, thanks



#83334 Looking for FASTA format of DNA of a gene amongst various species

Posted by humalog on 12 August 2010 - 03:30 PM in Bioinformatics and Biostatistics

Thank you Trof



#78148 Looking for FASTA format of DNA of a gene amongst various species

Posted by humalog on 05 July 2010 - 03:37 AM in Bioinformatics and Biostatistics

Thanks
Most of you thought I'm looking for amino acid sequence but some answers are helpful.

I wanted to ask: when I am in the EnsEMBL website and I search for a gene and get the results, how can I view the FASTA sequences of these?
thanks



#72916 Looking for FASTA format of DNA of a gene amongst various species

Posted by humalog on 27 May 2010 - 10:02 AM in Bioinformatics and Biostatistics

Hi

I need the DNA sequences (in FASTA format) of a specific gene (the tissue transglutaminase gene) for as many species as possible.

I've been able to find it for a few species, on the Entrez Gene database and on Entrez Nucleotide database.

Specifically the way I find these sequences is by searching for the gene name (TG2, which is the abbreviation for my gene of interest) in the search engines of these databases.

The problem is I only found the TG2 sequence for a handful of species (about 7), while the gene is supposed to be spread in almost all species.


Is there a more comprehensive way for finding the FASTA sequence of a gene throughout all species?


Thanks.



#71651 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 19 May 2010 - 08:56 AM in General Lab Techniques

Homebrew, thanks a lot.
Now I definitely got it.



#70778 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 13 May 2010 - 02:03 PM in General Lab Techniques

OK I think I understand now. The sybrsafe is what makes the DNA visible under UV. And the loading dye is just for making it visible under naked eye.

Thanks very much.
:P

Of course this means I'm doing something wrong with my PCR but I'll have to research it. Thanks again.



#70763 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 13 May 2010 - 12:40 PM in General Lab Techniques

To clarify, the DNA ladder readily has its own dye (from the moment of buying it). I don't add any dye to it.

I understood what the "stop solution" is, but I want to ask if this solution should also be visible under UV.

My stopping solution is this: http://www.promega.c...productleaf_602

Do you think it should also make the samples visible under UV?
(maybe it's a stupid question but I am relatively a beginner)



#70757 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 13 May 2010 - 12:12 PM in General Lab Techniques

Hi

I tried again and it still didn't work. This time I saw no bands at all (but still the ladder was visible clearly).

I have a new suspicion that I am doing something wrong after the PCR (or after the restriction digest- I use both these techniques with no good results).

Before I add the DNA PCR product (or the digested DNA in the case of r. digestion) I always add about 2ul of blue/orange dye 6x from Promega. Then I load samples in gel.

Today I saw that even though the bands were visible in the gel with naked eye (as orange or blue), when I put the gel under UV camera, the sample bands were not visible (only the ladder was).

So I wanted to ask you guys whether this blue/orange dye 6x is supposed to be used just for coloring the bands for observing during electrophoresis, or should it also make the bands visible under UV.

In case the first possibility is correct, then I guess I should also be adding a separate reagent to make the bands visible under UV. What would this be?



#70342 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 11 May 2010 - 09:11 AM in General Lab Techniques

Hi

Thanks for your responses.

Yes actually when I was saying "TE buffer" I meant to say "TAE buffer" and when I said "10ml of sybrsafe" I meant "10ul".

So you're right, I must be doing something wrong in my experiments and thanks for your suggestions I'll try adding more of DNA in each well next time.



#70165 Too much failure in electrophoresis, am I doing the gel correctly?

Posted by humalog on 10 May 2010 - 01:51 PM in General Lab Techniques

Hello.

I do experiments in a lab (I'm still a student) and I always do my agarose gel in a certain way. Most of the times however I don't get the expected bands revealed in the gel after electrophoresis. However I do get all the DNA ladder bands revealed.

I wanted to ask if there is a chance I might be doing something wrong when making up my gel. Or does the fact that the ladder is revealed mean that the gel is OK?

So I will explain how I do the gel for the electrophoresis and please let me know if it is wrong.

1) I add 980ml distilled water with 20ml TE of 50X concentration (I do this much so that I can keep some as stock).
2) I mix and add 100ml of this with 1g agarose powder and microwave this until all powder is dissolved.
3) I add 10ml of cybrsafe or however it is spelled.
4) I mix, pour it in the tray and let it dry for about 30 min.
5) I put the gel in the electrophoresis tank, and then fill the tank with the TE buffer I prepared at step 1.


So does my method seem right to you?
Thanks for listening to me and I look forward to hearing from you.




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