I have been more interested to read above-mentioned topic, give me your valuable answer with research articles.
I am working with Vibrio phage.
With Thanks and Regards,
-Stalin.n
Hi,
To rephrase your question how to screen out for GAL gene containing bacteriophage from the plagues.
Answer : PCR
From your isolated plagues, your need to extract enough DNA and to perform the PCR with Primer specific for full length GAL gene. The PCR amplificon corresponding to the size of GAL gene on EtBr gel electrophroesis is a indication of the presence of GAL in the your plague isolate.
Hence you have to get GAL sequence in NCBI data base and order an Primer .
I hope this answer was useful.
Rajkumar
I have to agree with the colleague Rajkumar Dhanaraju that from your isolated plagues, you need to extract enough DNA and to perform the PCR with Primer specific for full length GAL gene. The PCR amplificon corresponding to the size of GAL gene on EtBr gel electrophroesis is a indication of the presence of GAL in the your plague isolate. Igor Styriak
I agree with plaque-PCR, the trick is how to do it. I plated the phage-infected bacteria not in soft agar, but in soft agarose (0.5%). I picked plaques with a cut-off pipette-tip, and suspended them each into 20 uL TBE buffer, containing 0.25 u Agarase. I incubated at 42C for 4h, then used 1 uL as a template in a 25 ul volume of PCR. This liberates phages from the agarose-plugs, but still leaves them alive for further re-infection of bacterial cells. See my publication for details:
Article Bacteriophage recombineering in the lytic state using the la...
If the promoter of your GAL gene is active in E. coli and your host is LacZ- you could try an induction with lactose and screening replicas with X-GAL
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