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Asked 3rd Oct, 2012
  • Central Proteina Prima Company of Jakarta, Indonesia

How to design probe specific hybridize with a specific targeted gene in Southern blot method?

I use DIG-11-dUTP label probe with specific targeted gene to detect in host genome. Host genome have a huge sequence that to date no information in data base. I cut the genomic DNA by restriction enzymes but after southern blot process my specific gene in the host genome does not show the signal band. I use chemilluminence (SDP star) substrate for alkaline phospatase, I do not use radioactive label 32P. Could anyone give me more ideas with this case?
17th Oct, 2012
Meetul Kumar
Defence Research and Development Organisation
try temp 65 degree for hybridization. also the labelled probe must be single strand, use PCR high temp 95 degree. low stringy wash is a good option. check the pH of the buffers that you are using. most important.