About microscopic forms of life, including Bacteria, Archea, protozoans, algae and fungi. Topics relating to viruses, viroids and prions also belong here.
Actually, I want knockout eight genes.
And I transform 16 PCR products (eight of them are amplified using PKD3 as templete, and others are amplified using PKD4) to 16 competent cell. I did this at least 5 times. None of them seems work.
I did every thing I can do, such as using different L-ara concentration to induce RED recombination enzymes, using different PCR product concentration to transform competent cell, using different drug (kanamycin or chloramphenicol) concentration.
The results are still "I can get several dozen colonies from plate with kanamycin, but cann't get any colony from chloramphenicol.
" and "I cann't amplify anti-kanamycin gene from these anti-kanamycin strain."
I need suggestionssss......
OK, take your plasmid, which you use for transformation and try the PCR to see, whether your primers B work with your conditions
BTW in biology we use false positive, not fake
Cis or trans? That's what matters.
Your "BTW" is very constructive. Thank you, JackBean
You are the "JackBean" to me
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