Hi,
I just started using this strain of B.subtilis, and during transformation (Heat shock, natural competence), I noticed the negative and positive control plates looked exactly the same (I am using 50ug/ml kanamycin Lb plates for selection). Since this was my first time transforming this strain, I also used varying amounts of DNA (10, 100, 1000ng) as well as plating the transformation mix in various dilution factors (10, 100, 1000). The results were same in all dilution factors.
Initially I thought the plates were old and the kanamycin has degraded but I tried plating other bacteria (E.faecalis, WT B.subtilis, etc.) on it and it killed them all. Regardless, I made fresh 50ug/ml kanamycin Lb plates and tried plating B.subtilis WB800N and it actually grew.
Does anyone has any experience with this strain and if so, could anyone help? I recall reading some papers that used kanamycin for selection. The product listing also only mentioned having neomycin resistance, nothing about kanamycin. Did I miss something?
I have since learnt that the neomycin resistance gene codes for neomycin-kanamycin phosphotransferase, type II enzyme and confers resistance to aminoglycoside drugs such as neomycin and kanamycin-bacterial selection and G418-eukaryotic cell selection.
Article Synthetic neomycin-kanamycin phosphotransferase, type II cod...
Article Complete Genome Sequence of Bacillus subtilis Strain WB800N,...
I think it's a different type of enzyme that's conferring resistance but this paper seems to confirm the insertion of a kanamycin resistance gene in WB800N
Alexandra Johnson I emailed MoBiTec and they told me that they only inserted a neo resistance gene (which codes for that enzyme) into the "cm" gene disrupting it. I assume the "cm" they mentioned is the chloramphenicol resistance gene based on the article showing the complete genome.
But yea it pretty much confirmed that it has kanamycin resistance. Oh well.
- Badges
- Science topic
- Similar topics
- Physical Sciences
- Materials Science
- Plating