农业科学与技术（英文版）
農業科學與技術（英文版）
농업과학여기술（영문판）
AGRICULTURAL SCIENCE & TECHNOLOGY
2014年
5期
854-857
,共4页
大肠杆菌表达系统%毕赤酵母表达系统%植酸酶
大腸桿菌錶達繫統%畢赤酵母錶達繫統%植痠酶
대장간균표체계통%필적효모표체계통%식산매
Escherichia coli expression system%Pichia pastoris expression system%Phytase
[目的]研究外源基因在原核生物和真核生物中的表达。[方法]将带有目的基因的表达载体 pTYB2-WF转化大肠杆菌 ER2566，用 IPTG诱导植酸酶基因表达。用 SDS-PAGE验证植酸酶融合蛋白表达情况，并进一步对融合蛋白进行纯化。用?赤酵母表达系统表达植酸酶基因 phyA；构建酵母整合载体pPIC9K-phyA,转化?赤酵母 GS115，构建工程菌 GS115-pPIC9K-phyA。[结果]在甲醇的诱导下表达植酸酶，经酶活力的测定，其植酸酶活力达7.3μ/ml，构建了?赤酵母工程菌GS115-pPIC9K-phyA。[结论]甲醇酵母表达机制在分子生物学以及工业应用领域发挥作用。
[目的]研究外源基因在原覈生物和真覈生物中的錶達。[方法]將帶有目的基因的錶達載體 pTYB2-WF轉化大腸桿菌 ER2566，用 IPTG誘導植痠酶基因錶達。用 SDS-PAGE驗證植痠酶融閤蛋白錶達情況，併進一步對融閤蛋白進行純化。用?赤酵母錶達繫統錶達植痠酶基因 phyA；構建酵母整閤載體pPIC9K-phyA,轉化?赤酵母 GS115，構建工程菌 GS115-pPIC9K-phyA。[結果]在甲醇的誘導下錶達植痠酶，經酶活力的測定，其植痠酶活力達7.3μ/ml，構建瞭?赤酵母工程菌GS115-pPIC9K-phyA。[結論]甲醇酵母錶達機製在分子生物學以及工業應用領域髮揮作用。
[목적]연구외원기인재원핵생물화진핵생물중적표체。[방법]장대유목적기인적표체재체 pTYB2-WF전화대장간균 ER2566，용 IPTG유도식산매기인표체。용 SDS-PAGE험증식산매융합단백표체정황，병진일보대융합단백진행순화。용?적효모표체계통표체식산매기인 phyA；구건효모정합재체pPIC9K-phyA,전화?적효모 GS115，구건공정균 GS115-pPIC9K-phyA。[결과]재갑순적유도하표체식산매，경매활력적측정，기식산매활력체7.3μ/ml，구건료?적효모공정균GS115-pPIC9K-phyA。[결론]갑순효모표체궤제재분자생물학이급공업응용영역발휘작용。
Objective This study aimed to investigate the expression of exogenous gene in prokaryotes and eukaryotes. [Method] The expression vector pTYB2-WF harboring target gene was transformed into E. coli ER2566. IPTG was employed to induce the expression of phytase gene. The expression of phytase fusion protein was detected by SDS-PAGE, and the fusion protein was further purified. Phytase gene phyA was expressed in Pichia pastoris expression system. Yeast recombinant vector pPIC9K-phyA was constructed and transformed into P. pastoris GS115 to construct engineering strain GS115-pPIC9K-phyA. [Result] Phytase protein was ex-pressed under methanol induction. Enzyme activity assay indicated that the activity of phytase was 7.3 U/ml. P. pastoris engineering strain GS115-pPIC9K-phyA was successful y constructed. [Conclusion] Methanol yeast expression mechanisms play a certain role in molecular biology and industrial applications.
