Multiplexing first round PCR in BSP - Any experience? (Feb/14/2007 )
Good work Krumel,
can you tell us what protocol you used to make this multiplex i.e. what do you reccomend to those who want to do the same with regards to primer design and PCR set-up?
Hi Bram, if it is okay for you, I will just wait until Tuesday or Wednesday, when I got the sequencing results ...
Of course Krumel.
I had a go myself and it seems to work. I just took two primer sets which amplified regions on different chromosomes, with similar Tm and added them in the first round PCR. The second round I amplified the products per locus in different reactions and the results can be found below. Sample 1 did not work but the second one showed the desired bands. the difference between the two samples is the primer combinations and the type of DNA used. Sample 1 had digested and converted DNA while sample 2 had only converted DNA.
I am sure this would be a biotechniques paper!! As for acknowledgements? acknowledge the forum, it's a great place to share ideas and tips!
looking very promising Kruel. let's see what the sequencing tells you!
@Bram - cool - multiplexing the first round works for at least 4 genes (i.e. 8 primers). Thanks for replicating.
@Nick - good idea, acknowledging the paper
I will keep you up to date!
Was just wondering if there was any updates on the multiplex for BSP. Sounds like a great idea and if any papers have gone out on the topic could anyone share the references?
my results are promising but not yet definitive. I'll keep you informed as soon as a conclusion can be drawn.