目的　克隆酿酒酵母的蔗糖转换酶基因 (suc2 ) ,并定位突变酵母基因组中的 suc2基因以获得无蔗糖转换酶表达的酵母品系 .方法　用 PCR法从酵母 Y190基因组中扩增出suc2的成熟肽基因片段 ,克隆后测序 .将色氨酸合成酶 (TRP)基因片段插入 suc2基因中 ,构建成用于同源重组的 suc2基因定位突变载体 .导入酵母 Y190 ,用营养缺陷筛选出 suc2基因突变的克隆 .用 PCR扩增并克隆突变后的 suc2基因 ,用序列分析确定同源重组的发生 .结果　用 PCR从酵母基因组DNA中扩增出大小约 1.5 kb的 suc2基因片段 ,序列测定表明除 5 85位有一点突变 (AAC变为 AAT)外 ,所得 suc2基因与文献报道相同 ;构建的定位突变载体导入酵母细胞后 ,得到4株营养型符合的突变体 ;PCR表明这些突变体中均有 suc2基因的同源重组 ;取其中 1株的 PCR产物进行序列测定证实了同源重组的发生 .结论　克隆了正确的编码蔗糖转换酶成熟肽的 suc2基因片段 ;用同源重组方法在酵母 Y190的基因组中定位突变了 suc2基因 Objective To clone the suc2 convertase gene of Saccharomyces cerevisiae and locate the suc2 gene in the mutant yeast genome to obtain the yeast strain without sucrase converting enzyme.Methods The mature peptide gene of suc2 was amplified from the Y190 genome by PCR The sequence of the tryptophan synthase (TRP) gene was inserted into the suc2 gene to construct the mutated vector of suc2 gene for homologous recombination.After introducing into yeast Y190, the mutant with suc2 gene was screened out by auxotrophic factor PCR was used to amplify and clone the mutated suc2 gene, and sequence analysis was used to confirm the homologous recombination.Results The 1.5 kb fragment of suc2 gene was amplified by PCR from yeast genomic DNA, and the sequence analysis showed that except the 5 85 A little mutation (AAC changed to AAT), the obtained suc2 gene was reported in the literature; the constructed locus mutation vector was introduced into yeast cells to obtain four nutrient-matched mutants; PCR showed that these mutants have suc2 gene Homologous recombination, one of the PCR products was sequenced to confirm the occurrence of homologous recombination.Conclusion Cloning of the correct coding sugarcane Converting enzyme mature peptide suc2 gene fragment; by homologous recombination in the yeast genome method of positioning Y190 mutated gene suc2