XIE Baogui; LU Qiquan; RAO Yongbin; SUN Shujing; ZHENG Jingui Fujian Agricultural and Forestry University; Fuzhou; Fujian 350002; China

【中文摘要】 A human insulin gene was synthesized using PCR-based methodology, cloned into plasmid pUC18, and the sequence of the inserted fragment was confirmed. A Tremella fuciformis expression vector was constructed and used to transform T. fuciformis yeast-like conidia employing Restriction Enzyme-Mediated DNA Integration (REMI). A total of 21 antibiotic resistant colonies were selected at random of which 18 tested positive for β-glucuronidase (GUS) activity. Genomic DNA was extracted from 10 of the 21 p...更多ositive transformants and PCR performed using genomic DNA as the template and specific primers for amplifying the human insulin gene, the GUS gene and the Tnos sequence. PCR data showed that the expected fragment was amplified from all 10 isolates demonstrating that they were true human insulin gene transformants.  還原

【英文摘要】 A human insulin gene was synthesized using PCR-based methodology, cloned into plasmid pUC18, and the sequence of the inserted fragment was confirmed. A Tremella fuciformis expression vector was constructed and used to transform T. fuciformis yeast-like conidia employing Restriction Enzyme-Mediated DNA Integration (REMI). A total of 21 antibiotic resistant colonies were selected at random of which 18 tested positive for β-glucuronidase (GUS) activity. Genomic DNA was extracted from 10 of the 21 positive transformants and PCR performed using genomic DNA as the template and specific primers for amplifying the human insulin gene, the GUS gene and the Tnos sequence. PCR data showed that the expected fragment was amplified from all 10 isolates demonstrating that they were true human insulin gene transformants.

【英文關(guān)鍵詞】 Human insulin; Gene synthesis; Gene transformation; Tremella fuciformis; Yeast-like conidia
【基金】Sponsored by the National Natural Science Foundation ( No 30471210);; the Fujian Government Foundation(No WKL2005-2-009)

【文獻(xiàn)出處】 食用菌學(xué)報(bào),Acta Edulis Fungi,編輯部郵箱,2007年02期 【DOI】CNKI:SUN:SYJB.0.2007-02-001