need help: QUICK & DIRTY RT-PCR

HOFSTRA ANGELA 905-822-3520 Hofstra at aa.wl.com
Fri Aug 4 06:17:12 EST 1995


Hi all:

Does anybody knows a protocol that allows RT-PCR (of rather rare
messengers) to be carried out directly on CRUDE cell lysates ??
While for genomic PCR this is not a problem, i couldn't find such a "QUICK
& DIRTY" protocol that worked for RT-PCR. I need to directly RT-PCR
messengers out of many very small cell cultures (eg 24 well plates,
non-confluent), and I would like to circumvent the tedious pocedure of RNA
isolation from such small amount of cells. One other advantage of  RT-PCR
or PCR directly from lysates is the smaller sample-to-sample variation
introduced by any further steps of handling.
Thanks for any help and sugestions.

Sui Huang
Surgical Research
Children's Hospital
Boston, MA 02115
FAX: (617) 355-7043
TEL: (617) 355-5010
e-mail: SEHuang at aol.com


No affiliation to any company.

Amersham has a kit called the lysate mRNA capture kit for RT-PCR which I
believe will do exactly what you want.  In a slightly different vein Ambion has
a kit for doing RNase protection on cell lysate.  Hope this helps.

Angela





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