树状多节孢的双亲灭活原生质体融合  

INACTIVATED PARENTS PROTOPLAST FUSION OF NODULISPORIUM SYLVIFORME

作　　者：孙剑秋[1] 周东坡[1] 马玉超[1] 李晶[1] 解玉红[1] 张鹏[1] 平文祥[2] 

机构地区：[1]齐齐哈尔大学生命科学与工程学院,齐齐哈尔161006 [2]黑龙江大学生命科学学院,哈尔滨150080

出　　处：《菌物系统》2002年第3期430-436,共7页Mycosystema

基　　金：黑龙江省自然科学基金资助项目(93-C-11);黑龙江省"九五"重大攻关项目(G98C20-16-1)

摘　　要：以紫杉醇产生菌树状多节孢HQD33的诱发突变株UL50-6为出发菌株，将收集到的出发菌株UL50-6和UL40-19的菌丝体分别用pH5.5-6.0的0.7mol/L NaCl配制的3%溶壁酶、2%蜗牛酶、1%溶菌酶组成的复合酶系，30℃恒温酶解3-5h，制备原生质体。两菌株的原生质体经纯化后分别用热和紫外线灭活，其中UL50-6的原生质体在54℃热灭活5分钟，UL40-19的原生质体在30W紫外灯下，30cm，照射85秒进行紫外灭活，双亲株的原生质体存活再生率为零。同时对融合条件进行了初步探索，以含有Ca^2+和Gly的35%-40%的PEG作为融合剂，融合时间为20分钟时，融合率可以达到4.44×10^-2-6.92×10^-2。对融合株TPF-1与双亲株的形态学、可溶性蛋白、过氧化物同工酶进行分析，确证其为双亲株的融合子。The mutant UL50-6 and UL40-19 of the fungus producing Taxol-Nodulisporium sylviforme HQD33 is digested by compound enzyme consisting of 0.7mol/L NaCl, 3% lywallzyme, 2% snailase and 1% lysozyme for 3~5 hours at the condition of 30℃ and pH5.5~6.0. The protoplasts of UL50-6 and UL40-19 are obtained. The regeneration frequency of the parents protoplast is zero when the protoplast of the mutant Ul50-6 is heat-inactivated at 54℃ for 5 minutes and the protoplast of the mutant UL40-19 is UV-inactivated at 30w, 30cm ,85 seconds. The fusion conditions are probed initially. When the inactivated protoplast fusion is induced by 35%~40% PEG containing Ca2+ and Gly for 20 minutes, fusion frequency may come up to 4.44?0-2~6.92?0-2. The fusant TPF-1 is compared with the parents in terms of morphology, soluble protein and isozyme graph and so on. The comparison shows that TPF-1 is the fusant of the parents really.

关 键 词：树状多节孢 原生质体 融合 双亲灭活 

分 类 号：Q813.2[生物学—生物工程]