质粒DNA对雪白链霉菌原生质体的高频转化

High frequency transformation of Streptomyces niveus protoplasts by plasmid DNA.

作者信息

Hussain H A, Ritchie D A

机构信息

Department of Genetics and Microbiology, University of Liverpool, UK.

出版信息

J Appl Bacteriol. 1991 Nov;71(5):422-7. doi: 10.1111/j.1365-2672.1991.tb03811.x.

DOI:10.1111/j.1365-2672.1991.tb03811.x

PMID:1761435

Abstract

A procedure has been developed for transforming protoplasts of the novobiocin producing strain Streptomyces niveus at high frequency. This required the isolation of strains LH13 and LH20 defective in DNA restriction from the wild type (ATCC 19793) which is transformed at very low frequencies. The LH13 and LH20 derivatives were obtained by curing pIJ702 DNA from the few S. niveus transformed protoplasts obtained by transformation of the wild type with high concentrations of pIJ702 DNA. Protoplasts of S. niveus strains LH13 and LH20 produced about 10(6) transformants/micrograms DNA with modified pIJ702 DNA derived by replication in S. niveus. Unmodified DNA (derived from replication in S: lividans) from a series of pIJ101, SCP2 and pSN2-based derivatives, gave transformation frequencies in the range of 10(2)-10(3) transformants/micrograms DNA. Optimal conditions for the formation and transformation of S. niveus protoplasts are described.

摘要

已开发出一种程序，可高频转化新霉素产生菌雪白链霉菌的原生质体。这需要从野生型（ATCC 19793）中分离出DNA限制缺陷型菌株LH13和LH20，野生型的转化频率非常低。LH13和LH20衍生物是通过从用高浓度pIJ702 DNA转化野生型获得的少数雪白链霉菌转化原生质体中去除pIJ702 DNA而得到的。雪白链霉菌菌株LH13和LH20的原生质体用在雪白链霉菌中复制得到的修饰pIJ702 DNA产生约10(6)个转化子/微克DNA。来自一系列基于pIJ101、SCP2和pSN2的衍生物的未修饰DNA（在变铅青链霉菌中复制得到），其转化频率在10(2)-10(3)个转化子/微克DNA范围内。描述了雪白链霉菌原生质体形成和转化的最佳条件。