Hello everyone,
How can a researcher design the primers for a gene of interest using Uniprot.org.A researcher searches the gene of interest in Uniprot.org. in which he gets the gene name alongwith a bunch of detailed information and the sequences of that gene. The sequence shown is usually like for eg:
10 20 30 40 50 MSSSDEETLS ERSCRSERSC RSERSYRSER SGSLSPCPPG DTLPWNLPLH 60 70 80 90 100 EQKKRKSQDS VLDPAERAVV RVADERDRVQ KKTFTKWVNK HLMKVRKHIN 110 120 130 140 150 DLYEDLRDGH NLISLLEVLS GIKLPREKGR MRFHRLQNVQ IALDFLKQRQ and so on.
The options available are FASTA,Blast etc in Uniprot.org.
I would like to know,how can a researcher design the primers for this gene of interest using these sequences?
Note: Kindly attach the links whatsoever you have and you can.
Thanking you in advance,
Hi Mr Rouf
I assumed you want to design the primer for the gene coding sequence.
In this case you can search the gene in the genebank based on the UniProt Gene name and organism, find the mRNA sequence then copy its Genebank Accession.
You can use PrimerBLAST in this case:
https://www.ncbi.nlm.nih.gov/tools/primer-blast/index.cgi?LINK_LOC=BlastHome
Open the site and paste the accession number in the entry box.
The other way is by translating the amino acid sequence from the UniProt using related software e.g BioEdit, MEGA, or simply go to online translator sites such as
https://www.ebi.ac.uk/Tools/st/emboss_backtranseq/
https://www.bioinformatics.org/sms2/rev_trans.html
It is also possible to upload the FASTA format (nucleotide sequence) or copy-paste the translated sequence in the input box at the PrimerBLAST
By using PrimerBLAST you can also check the specificity of the primer by compare it to the other sequence available in the data base.
You can also use Primer3 program : http://primer3.ut.ee/ to desain the primer. Just paste the sequence in the input box an define the primer condition based on your interest
Good Luck
Find your gene, intron/exon structure and surrounding sequences using this tool: http://genome.ucsc.edu/
Just pop your gene sequence into BLAT for the organism you want and go.
I recommend you the easiest way to get probed primers by searching in published papers the sequences they used or searching in primers databases such as primerdepot or primerbank.
I hope this link helps you.
https://pga.mgh.harvard.edu/primerbank/
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