■ 基本信息
別名: | pET32a |
啟動子: | T7 |
復制子: | pBR322 |
終止子: | T7 terminator |
質(zhì)粒分類: | 大腸系列質(zhì)粒；大腸表達質(zhì)粒；pET系列質(zhì)粒 |
質(zhì)粒大小: | 5900bp |
質(zhì)粒標簽: | N-Trx，N-6×His，N-thrombin，N-S，N-enterokinase，C-6×His |
原核抗性: | Amp |
克隆菌株: | DH5a |
培養(yǎng)條件: | 37℃，有氧，LB |
表達宿主: | BL21(DE3)等大腸桿菌 |
誘導方式: | IPTG或乳糖及其類似物 |
5'測序引物: | T7：TAATACGACTCACTATAGGG |
3'測序引物: | T7-ter：TGCTAGTTATTGCTCAGCGG |
■ 質(zhì)粒屬性
質(zhì)粒宿主: | 大腸桿菌 |
質(zhì)粒用途: | 蛋白表達 |
片段類型: | ORF
|
片段物種: | 空載體
|
原核抗性: | Amp |
真核抗性: |
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熒光標記: |
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■ 質(zhì)粒簡介
pET-32a質(zhì)粒是一種原核表達載體，C端含有一個6×His標簽，N端含有一個thrombin位點、6×His標簽、TrXA位點、Ek位點和S標簽。該質(zhì)粒含有多個常用的酶切位點，便于不同基因克隆。表達由宿主細胞提供的T7 RNA聚合酶誘導，目的基因被克隆到質(zhì)粒載體上，受噬菌體強轉(zhuǎn)錄及翻譯信號控制。
pET系統(tǒng)是有史以來在大腸桿菌中表達重組蛋白的功能最強大的系統(tǒng)，也是現(xiàn)今原核表達方面使用最廣泛的系統(tǒng)。該系列質(zhì)粒能很容易的通過降低誘導物的濃度來削弱蛋白表達。在非誘導條件下，可以使目的基因處于完全沉默狀態(tài)而不轉(zhuǎn)錄。
The pET-32 series is designed for cloning and high-level expression of peptide sequences fused with the 109aa Trx?Tag? thioredoxin protein (1). Cloning sites are available for producing fusion proteins also containing cleavable His?Tag? and S?Tag? sequences for detection and purification. Unique sites are shown on the circle map. Note that the sequence is numbered by the pBR322 convention, so the T7 expression region is reversed on the circle map. The cloning/expression region of the coding strand transcribed by T7 RNA polymerase is shown below. The f1 origin is oriented so that infection with helper phage will produce virions containing single-stranded DNA that corresponds to the coding strand. Therefore, single-stranded sequencing should be performed using the T7 terminator primer.
質(zhì)粒只保證關(guān)鍵序列正確，不保證表達效果。
■ 質(zhì)粒圖譜
■ 質(zhì)粒序列
質(zhì)粒序列請下載：
ZK170pET-32a大腸表達質(zhì)粒.txt
