Two Evidences Proving the Activity of YcaI (com2 homologue)

Firstly I provide the two observations which strongly suggest that ycaI dependent transformation actually also participate in our transformation system.

1. 24 h after replica, only the wild type without antibiotic stimulation can still produce transformants. The ycaI mutant, the wild type and the ycaI mutant pretreated with low concentration of antibiotics gives few (most are 0) transformants at this period. It seems that antibiotics induce transformation occurring earlier. This might be true for the ycaI-independent pathway, but another evidence reveal that the two mechanism seems not be the same.

2. When I do the replica experiment, I replicate the lawn from LA plates to two plates, one contains high concentration of Ampicillin and the other contains high concentration of spectinomycin. In ycaI mutant, comparing the LA-Amp plate and LA-Spc plate, the layouts of transformants are almost the same. However, for the wild type, the layouts of transformants on LA-Amp and LA-Spc are completely different.

How to explain this phenomenon. Remember my last post, since transformation occurrs only on plates we can observe transformants in situ. Two possibilities might be able to explain this phenomenon: 1. transformants moved. 2. the two transformantion mechanism are different, so the transformants are derived from different types of competent cells. So they are intrincically different. If first possibility is true, how cells move? Does this means that type IV pili is active at this moment and largely enhanced the mobility of transformants cells? One thing is clear that ycaI-dependent transformation behavior is slower than that of ycaI-independent transformation behavior (the first evidence I posted here). If the second possibility is true, that is the transformants observed on ampicillin plate and spectinomycin plate derived from different cells. Why one type of cells can grow on ampicillin plate and the other grow only on spectinomycin? Since they have finished transformation process and both of the amp resistance gene and spc resistance genes have entered into cells, they should be able to resistant to both of the two type of transformants. The situation is similar to the paper published more than 20 years ago. The author said that they can not explain the phenomenon, that transformation of ampicillin marker in ycaI mutant with a plasmid containing USS is much lower than transformation of ampicillin marker in the wild type cell with the same plasmid without USS. But for nov resistance marker, the former for of transformation was much stronger than the latter one. Both of my results and the author's support that once cells decide to adpot one type of transformation, the other type is closed.