To study the conjugal effect of P3 fragment of Streptomyces linear plasmid SCP1
| Autor: | Han-Rong Wu, 吳翰融 |
|---|---|
| Rok vydání: | 2018 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 106 Gram-positive bacteria Streptomyces is different from other bacteria keeping circular chromosome, the topology of their chromosomes are linear. The conjugal circular or linear plasmids are usually observed in Streptomyces, but the detail mechanism is not clear. Current studies have shown the circular plasmids performed DNA transfer into recipient in double-stranded format, and the TraB recognize a cis-acting locus of transfer (clt) which give the transfer ability of linking DNA fragment. In conjugal linear plasmid part, clt is not yet identified. Our previous studies have found a clt candidate of SCP1 linear plasmid in an internal 3.2 kb DNA fragment. This study was continued to narrow down the clt. The 3.2 kb DNA fragment was separated into three overlapping fragments, then inserted them into a constructed linear plasmid without transfer ability for test. The plasmids carrying tsr (thiostrepton resistant gene) were transfer into S. lividans 3200 to be a donor strain, then mates with recipient strain S. lividans TK54. The plasmids transfer efficiency was counted by thior recombinants. The results showed that the transfer efficiency of those three DNA fragments were 100 times less than the intact 3.2 kb DNA fragment, suggested the clt of SCP1 was more dispersive than those in circular plasmids. In further, several constructs with deletion from both sides and internal regions of the 3.2 kb were examined, the results showed the 300 bp of both sides and the internal 281 bp were not required for the clt function. These results suggested the clt should be separated to two parts and require further experiments to understand the conjugal mechanism of SCP1 linear plasmids. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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