handling cDNA - care in handling (Nov/22/2005 )

can someone kindly tell me the steps and care taken in handling cDNA? i bought human lung cDNA and tried to amplify a family of related genes...

i manage to get what i want at first... but now, nothing seems to work...

i have changed all my pcr reaction mix [primers, dntp, buffer, supertaq and h20]

so my best guess is, i have damaged my cDNA:(

how how?

i thaw on ice... and flick vial a few times and 'whip' down... will this damage the cDNA?

I would think perhaps nuclease contamination

multiple freeze/thaw cycles are bad juju. whenever possible, make little aliquots

for good comprehensive info on cDNA handling (if you did not get product information from the cDNA vendor) try these websites: Ambion, Invitrogen, Promega

or do a Google for "cDNA handling"