| Summary: | 碩士 === 國立臺北科技大學 === 生物科技研究所 === 99 === Streptomyces lividans have a 50 kb SLP2 linear plasmid. The linear plasmid can be transfer to other Streptomyces during the conjugation. The current established mechanism of conjugation is mostly based on circular F plasmid in E. coli system. But that one-way, single-strand DNA transfer mechanism couldn’t be applied to linear plasmid conjugal transfer of Streptomyces. Our lab target on locating the replication origin (oriT) by identifying the clt (cis-acting locus of transfer) region of S. lividans SLP2 plasmid. Our previous study showed that non-transferrable linear plasmid carring 2 kb SLP2 clt fragment resulted in successful transfer to recipient. The data showed that the oriT is located in the 2 kb region. But yet to confirmed the DNA transfer during conjugation start from this region so this fragment is temporarily named clt.
Thesis focuses narrow down the 2 kb clt containing region. The non-transferrable homologous plasmids with different fragment of the 2 kb clt containing region and
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observing with plasmid with small clt fragment that can be transferred to the recipient under linear plasmid SLP2 conjugation background. By this strategy, I found the clt is a fragment of length about 450 bp. In many studies, tra had been reported as a main factor that helps plasmid transfer. However, overexpression of tra resulted in cytotoxicity. Thus, kil-kor system can be found in most of the Streptomyces to control tra expression. In this study, recombinant plasmid with tra not causing observable cell death. This result suggests that SLP2 tra might under regulation of gemonic kil-kor system or the regulatory mechanism is different from the general kil-kor/tra system.
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