Protoplast Isolation and Genetic Transformation of Polyporus brumalis

Abstract

This experiment was undertaken to investigate proper conditions for protoplast isolation and genetic transformation of the white rot fungi, Polyporus brumalis. The protoplasts were formed from mycelia at a frequency of 1 × 10 7 /ml with 0.5% Usukizyme. The transformation vector (pHYgpt) was constructed using hygromycin resistance gene (hph) for the selectable maker. The yield was 100-160 transformants/μg DNA in a transformation mediated by 40% polyethylene glycol solution with aurintricarboxylic acid, heparin and supermidine. The genomic integration of the pHYgpt was confirmed by hph-specific PCR and the expected amplified band appeared only in the transformants. These results could be an efficient tool in gene engineering of the genus polyporus. et al., 1993;Hatakka, 1994). 리그닌 분해효소는 기질특이성이 낮기 때문에 리그닌 외에 염료나 내분비계 장애물질 등 난분해성 유해화합물을 분해할 수 있으며, 분해과정이 세포외 에서 이루어지기 때문에 고농도의 독성화합물과 불용성화합물 의 분해도 가능하다 (Pasti-Grigsby et al., 1992;Barr and Aust, 1994;Pointing, 2001;Shin et al., 2007;Hwang et al., 2008