Sometimes when I am looking for some microorganisms primer genes in the web I found some of them, but have been used with different DNA organisms, can I use the same primers with different organisms?
Hello Mohammad,
it depence on what are you looking for and how specific are your primers. If they are very specific, probably they don´t work with any other specie. But if they have been designed from a very conservative sequence, could be they worked with other species. What are you looking for?
HI, you can use degenerate primers. But it's also tricky! David is right: It depends what you are looking for.
http://hcgs.unh.edu/protocol/basic/pcrdegenpri.html
Cheers, Nadine
Thank you all for response , actually I am looking for metallothionien in filamentous fungi ,I found some primers used in Daphnia pulex, can I used the same primers
The specificity of the primer designed also depends on its 3 ' side ( particularly first three bases), if your target sequences and your primers 3' side are maching then i suppose it should work.
Dear Mohammad lb. Khalil,
If the region of interest are similar for methallothionein gene in two different organims it would work, with some limitations (I agree with Naveed comment). If you have already the primers you can try to do some basic alignment. But if not, do not waste your time. The PrimerQuest tool is extremely easy to use for primer design (http://www.idtdna.com/primerquest/Home/Index)
Good luck!
Do you know primer blastn from ncbi? There you can paste your primer sequence and even the name of the microorganisms. But you can Start a search without Species limitation so you will get an alignment with all sequences published in ncbi. Check out the 3' ends oft the primers - no missmatches for better hybridisation
You can use ClustalW for multiple sequence alignment and if there is a descent % (>95%) match between gene sequences of two organisms you can try the primer pairs designed against a specific region of target sequence only if the nucleotide BLAST analysis come up with target gene name specific for the test organisms.
Using same primer for a gene in different organisms depends on the sequence conserve of a region from which the primer design. If sequence is conserved you can use same the primer for all organisms, else used specific primer for different organisms provided its gene sequence is known else design degenerated primers.
Depends on the sequence similarity. However, even though the primers might be specific for one particular species and you find that they might work with another species, you then have to worry about permiscuity of the primers in the different species genome. You may amplify un wanted amplicons that could migrate with your GOI and confound your study. You can easily just design the primers if you want by using NCBI primer blast or design them by eye. Look for 20-23mers that are ~50% GC rich and as mentioned above make sure the 3' ends are clamped down well; you can achieve this nicely if the primers end in a C or G.
good luck Mohammad!
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