rp-pcr - (Mar/31/2006 )

hi there

i need some help for re-pcr when i want to increase the cycling for the same product after one day

the bands not became thick means not happen pcr i don't know the reason

thank u

-maher-

rp-pcr? re-pcr?

would you please post more details of what you have done, and what you are trying to accomplish, so that we may help you?

-aimikins-

i do pcr for MDA prodect (Library) after i do on 25 cycle i see weak band

when i want to increase the cycle after one day i have the same band

thats mean the pcr not happen

i hope you help me

-maher-

so if I understand you correctly, you want to increase your yield...what have you tried so far? have you only tried to increase cycle number?

if you are limited by one of your ingredients, that will not improve your yield. perhaps trying a little more template will help?

-aimikins-

You want to re(peat) your PCR right? The bands are faint even after you increase the number of cycles right? You may want to try to increase the amount of sample you are using for your PCR, your PCR reagants, mix your reagants well, spin down the tubes before you place them in the PCR machine, or is it that you miscalculated you're primers concentration?

-moonstar-

QUOTE (moonstar @ May 3 2006, 04:47 AM)

hello everyone.
if i am right maher is doing a secondry pcr with using primary pcr product as a template to increase
i think u should try to change template conc. may b u should dilute it bacause sometimes in secondry pcr we dont get teh bands because in primary pcr product conc will b high, though we cant see nay band but still we hav to use diluted template which shows beautiful bands . so u try with various conc.
all the best
priya

-priya-