- I isolated some Gram-negative isolates that appear high resistance to most antibiotics. I want to do multiplex PCR to isolates for beta-lactam gene resistance. are there any recommendations to choose between mix of genes, there is any limit of primers in the same mix. how I choose the best mix of primers in the same mix, is there any recommendation to most recent genes needing study, also to help me publishing my paper speedily.
- Thanks in advance for every one
Dear Samah:
You can take at least 2 primers for each family of Major (TEM, SHV, CTX-M, and OXA) and Minor ( IMP VIM, NDM, KPC, etc...) beta-lactamase families, in addition to AmpC β-lactamases.
Regards
Dear Samah:
You can take at least 2 primers for each family of Major (TEM, SHV, CTX-M, and OXA) and Minor ( IMP VIM, NDM, KPC, etc...) beta-lactamase families, in addition to AmpC β-lactamases.
Regards
Make a list of resistance genes of all minor and major beta-lactamese families.
make multiplex Primers designing. and guide line for multiplex PCR conc. and temp. see below articles
Article MPprimer: A program for reliable multiplex PCR primer design
https://www.neb.com/protocols/2012/09/13/multiplex-pcr-guidelines-for-multiplex-pcr-5x-master-mix-m0284
Dear
It's not necessary to test all bêta-lactamase genes. Befor that you should determine the phenotype : carbapenemase (MBL, KPC, ...), pAmpC, ESBL, ... When you have done this step, it's then simple to choose your primers. If you need help, please feel free to contact me in private.
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