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基因的克隆與表達(dá)服務(wù)|實(shí)驗(yàn)技術(shù)服務(wù)

時(shí)間:2015/7/1閱讀:248
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關(guān)鍵詞:基因的克隆與表達(dá)服務(wù)|實(shí)驗(yàn)技術(shù)服務(wù)
簡介:世界*品牌基因的克隆與表達(dá)服務(wù)|實(shí)驗(yàn)技術(shù)服務(wù)原裝,,質(zhì)量保證,*,。
基因的克隆與表達(dá)服務(wù)|實(shí)驗(yàn)技術(shù)服務(wù)——pCzn1質(zhì)粒+Arctic Express宿主菌低溫表達(dá)體系,。
我們具備豐富的蛋白表達(dá)設(shè)計(jì)經(jīng)驗(yàn)及實(shí)驗(yàn)操作技巧,承諾客戶不成功不收費(fèi),。我們所有的實(shí)驗(yàn)數(shù)據(jù)真實(shí)可信,,我們提供原始的表達(dá)菌株和克隆質(zhì)粒,客戶可以依據(jù)我們的實(shí)驗(yàn)報(bào)告重復(fù)我們的實(shí)驗(yàn)結(jié)果,。
產(chǎn)品品牌:   http://www.,。。com/goodsid/fenleiyi/2868603/1.html    
測序?qū)嶒?yàn)流程:
從不同的重組DNA分子獲得的轉(zhuǎn)化子中鑒定出含有目的基因的轉(zhuǎn)化子即陽性克隆的過程就是篩選,。目前發(fā)展起來的成熟篩選方法如下:
(一)插入失活法
外源DNA段插入到位于篩選標(biāo)記基因(抗生素基因或β-半乳糖苷酶基因)的多克隆位點(diǎn)后,,會(huì)造成標(biāo)記基因失活,表現(xiàn)出轉(zhuǎn)化子相應(yīng)的抗生素抗性消失或轉(zhuǎn)化子顏色改變,,通過這些可以初步鑒定出轉(zhuǎn)化子是重組子或非重組子,。目前常用的是β-半乳糖苷酶顯色法即藍(lán)白篩選法。
(二)PCR篩選和限制酶酶切法
提取轉(zhuǎn)化子中的重組DNA分子作模板,,根據(jù)目的基因已知的兩端序列設(shè)計(jì)特異引物,,通過PCR技術(shù)篩選陽性克隆。PCR法篩選出的陽性克隆,,用限制性內(nèi)切酶酶切法進(jìn)一步鑒定插入片段的大小,。
(三)核酸分子雜交法
制備目的基因特異的核酸探針,通過核酸分子雜交法從眾多的轉(zhuǎn)化子中篩選目的克隆,。目的基因特異的核酸探針可以是已獲得的部分目的基因片段,或目的基因表達(dá)蛋白的部分序列反推得到的一群寡聚核苷酸,或其它物種的同源基因,。
(四)免疫學(xué)篩選法
獲得目的基因表達(dá)的蛋白抗體,就可以采用免疫學(xué)篩選法獲得目的基因克隆,。這些抗體即可是從生物本身純化出目的基因表達(dá)蛋白抗體,,也可從目的基因部分ORF片段克隆在表達(dá)載體中獲得表達(dá)蛋白的抗體。
奇妙的克隆克隆技術(shù)已展示出廣闊的應(yīng)用前景,,概括起來大致有以下四個(gè)方面:
(1)培育優(yōu)良畜種和生產(chǎn)實(shí)驗(yàn)動(dòng)物,;
(2)生產(chǎn)轉(zhuǎn)基因動(dòng)物,;
(3)生產(chǎn)人胚胎干細(xì)胞用于細(xì)胞和組織替代療法;
(4)復(fù)制瀕危的動(dòng)物物種,,保存和傳播動(dòng)物物種資源,。
表達(dá)策略:1 反向遺傳學(xué):克隆的CDS可以亞克隆到特定的表達(dá)載體,并轉(zhuǎn)基因到特定的物種(如果本物種的轉(zhuǎn)基因體系尚未建立或很困難,,可以先轉(zhuǎn)入一些模式生物),,按需要在各種啟動(dòng)子的驅(qū)動(dòng)下在生物體內(nèi)表達(dá),觀察表型,;同時(shí)可以根據(jù)該CDS序列,,設(shè)計(jì)RNAi載體,轉(zhuǎn)基因,,觀察表型,。如果需要,可以克隆該基因本身的啟動(dòng)子,,然后讓其驅(qū)動(dòng)該基因或RNAi片斷表達(dá),。
2 體外表達(dá):可以利用大腸桿菌、酵母系統(tǒng),,在體外做一些原核或真核的表達(dá),。用以研究酶活,制備抗體等,。

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