Has anyone used standard Polymerase in a long-RT-PCR?

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E. Rasp

Have you tested your cDNA (like with referencegene)? Quality of your RNA?

Up to 1Kb is possible. Maybe increase input Taq a little, but don't overdo. Try to increase the extensiontime. Also important is MgCl2-concentration. And sometimes standard Taq just won't give the results you want. Taq can be expensive, but when trying over and over again with no results is also expensive. In a normal DNA-PCR we got very good results (1700 bp) with QiaGen Hot Start Taq DNA polymerase.

Hung King Tiong

Are these primers binding to the exon regions of your gene mRNA cDNA of study?

If the primers are binding to the exon regions, then use 1 min extension time to produce 1kb amplicon.