PCR Using T7 and SP6 Primers

[Paul Fisher]

I've always used custom designed M13 forward and reverse sequencing
primers that anneal at 65 C . Now I'm using different primers designed
by someone else that anneal at 55 C, which also works fine. You can
design or purchase them in bulk and use them permanently. I think
designing primer sets is cheaper.

On 30 Jul 1997 19:24:47 -0700, ddsyr at NS.EAST.CN.NET (ma) wrote:

>Dear Netters:
>I want to check the insert in plasmid (pGEM), so I plan to use a universe primers, 
>say T7(5'AAT ACG ACT CAC TAT AG 3') and SP6(5' ATT TAG GTG ACA CTA TAG 3'), which 
>are also used in DNA sequence. But I find the Tm of these two primers are very low, 
>only 42-45. So I guess this may influence these PCR efficiency. Hope the experts 
>give me their experiences and suggestions. Thanks
>Ma
>