CAJ | 학술논문

为了克隆鹅生长激素基因并表达其重组蛋白质，采集生长期鹅垂体组织，并利用TRIzo1快速提取的总RNA为模板，反转录为cDNA.根据鹅生长激素基因编码的成熟肽序列(GenBank号：AY149895.2)设计1对引物，分别在上、下游引物的5忆端引入Nhe I和Hind III酶切位点.经反转录扩增获得鹅生长激素基因的编码的成熟肽全序列.通过双酶切和连接将鹅生长激素编码区插入原核表达载体pRSET-A的Nhe I和Hind III位点之间，构建重组表达质粒pRSET-gGH并转化大肠杆菌表达菌株BL21(DE3).转化的菌株经IPTG诱导后表达重组鹅生长激素蛋白质，分子量约为2.93×104.经过DEAE-650M弱阴离子交换树脂纯化获得较高纯度的重组鹅生长激素蛋白质.
위료극륭아생장격소기인병표체기중조단백질，채집생장기아수체조직，병이용TRIzo1쾌속제취적총RNA위모판，반전록위cDNA.근거아생장격소기인편마적성숙태서렬(GenBank호：AY149895.2)설계1대인물，분별재상、하유인물적5억단인입Nhe I화Hind III매절위점.경반전록확증획득아생장격소기인적편마적성숙태전서렬.통과쌍매절화련접장아생장격소편마구삽입원핵표체재체pRSET-A적Nhe I화Hind III위점지간，구건중조표체질립pRSET-gGH병전화대장간균표체균주BL21(DE3).전화적균주경IPTG유도후표체중조아생장격소단백질，분자량약위2.93×104.경과DEAE-650M약음리자교환수지순화획득교고순도적중조아생장격소단백질.
Tota1 RNA of goose growth hormone gene extracted from goose pituitary tissue with TRIzo1 reagent were used as temp1ate for reverse transcription of the first strand cDNA and a pair of primers was designed based on the pub1ished goose growth hormone gene sequence fragment (GenBank No. AY149895. 2), in which, two restriction enzyme sites Nhe I and Hind III were introduced into the 5’end of both upstream and downstream primers respective1y. Goose growth hormone gene coding sequence was amp1ified by PCR, and the amp1ified mature peptide sequence was inserted into the Nhe I and Hind III sites of the expression vector pRSET-A to generate the recombinant expression p1asmid pRSET-gGH which was transformed into Escherichia coli BL21(DE3) afterwards. The transformed bacterium was induced with IPTG to express the recombinant protein with a mo1ecu1ar mass of 2. 93×104. The high purity recombinant goose growth hormone was achieved using DEAE-650M weak anion exchange resin.