Hi,
I was just wondering if this will affect my downstream work. I've cloned my in situ probe into plasmid that has both T7 and SP6 promoters on each end and have done the in vitro transcription using the respective RNA polymerases. I've digested the resulting RNA/plasmid DNA mix with DNAse I according to Qiagen (incubate for 10 minutes at room temp) and have cleaned up using column based methods... Ran them on normal 1% agarose gel and have found that there is a faint band that corresponds to undigested plasmid DNA. Will this affect my in situs? Or should I remake the probes or redigest the mix again?
Help is greatly appreciated!
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