Construction of Expression Vector for High copy number Integration of the Methylotrophic Yeast Hansenula polymorpha
甲醇酵母表达系统高拷贝数整合型表达载体的构建

Based on the YIp type expression vector pJF5 1 of the methylotrophic yeast Hansenula polymorpha , a novel vector, pMIRH , for high copy number integration was constructed, which contains a random sequence from the ribosomal DNA of the host chromosome, a defficient maker gene Hp leu 2d derivated from the wild gene with a partial upstream sequence deleted and a G418 resistance gene ( neo ) controled by the SV40 early promotor. Results from experiments of transformation, screening and Dot and Southern hybridization showed that pMIRH can result in transformants with high copy number vectors, which can be screened by a secondary G418 resistance selection, and these vectors were integrated into the chromosome in a tandem arrangement.