I have a strain adapted to growing at higher temperature (47 degrees), when a plasmid was introduced for protein expression, it failed to grow, almost at all at that temperature it was previously adapted even without induction, but perfectly grew at it's normal growing temperature conditions (30 degrees).
what would be the cause? any solution?
Hello, adaptation of grow culture at high temperature mean that during adaptation you have same changes in DNA which give you adaptation. As we know adaptation process is mostly connected with plasmid DNA within cell. So by transformation you introduce another plasmid into cell and in this case you have at list two plasmids in one cell. So may be new plasmid inhibit the expression of second which is responsible for growth at high temperatures.
If you construct plasmid harboring insert A and insert B by using the same vector backbone ( same origin of replication). Then, transform two plasmid into same host. Plasmid will soomly loss. You can get more detail in website: https://blog.addgene.org/plasmid-101-origin-of-replication
Good luck!
You may be stressing the bacteria excessively. If it is temperature adapted to 47, it might be just on the edge of being able to grow. Adding a stress of an antibiotic selection might be too much. Try switching to 47 without using antibiotic in the media.
Hello, adaptation of grow culture at high temperature mean that during adaptation you have same changes in DNA which give you adaptation. As we know adaptation process is mostly connected with plasmid DNA within cell. So by transformation you introduce another plasmid into cell and in this case you have at list two plasmids in one cell. So may be new plasmid inhibit the expression of second which is responsible for growth at high temperatures.
If you construct plasmid harboring insert A and insert B by using the same vector backbone ( same origin of replication). Then, transform two plasmid into same host. Plasmid will soomly loss. You can get more detail in website: https://blog.addgene.org/plasmid-101-origin-of-replication
Good luck!
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