■ 基本信息
復(fù)制子: | pUC |
終止子: | SV40 poly(A) signal |
質(zhì)粒分類: | 哺乳系列質(zhì)粒；哺乳熒光質(zhì)粒；哺乳綠色質(zhì)粒 |
質(zhì)粒大小: | 4150bp |
質(zhì)粒標(biāo)簽: | C-AcGFP1 |
原核抗性: | Kan |
真核抗性: | G418 |
克隆菌株: | DH5a |
培養(yǎng)條件: | 37度 |
表達(dá)宿主: | 293T等哺乳細(xì)胞 |
誘導(dǎo)方式: | 無(wú)須誘導(dǎo)，瞬時(shí)表達(dá) |
5'測(cè)序引物: | AcGFP-1-F(GCGTCGATTTTTGTGATGCTC) |
3'測(cè)序引物: | Sv40-polyA-R（GAAATTTGTGATGCTATTGC） |
備注: | 哺乳細(xì)胞啟動(dòng)子檢測(cè)載體 |
■ 質(zhì)粒屬性
質(zhì)粒宿主: | 哺乳細(xì)胞 |
質(zhì)粒用途: | 信號(hào)報(bào)告 |
片段類型: | Promoter
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片段物種: |
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原核抗性: | Kan |
真核抗性: | G418
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熒光標(biāo)記: |
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■ 質(zhì)粒簡(jiǎn)介
pAcGFP1-1質(zhì)粒是 AcGFP的衍生物，來(lái)自Aequorea coerulescens。 AcGFP1已經(jīng)優(yōu)化了更亮的熒光。（激發(fā)最大值= 475nm;發(fā)射最大值= 505nm）AcGFP1基因的編碼序列含有沉默堿基變化，其對(duì)應(yīng)于人類密碼子使用偏好。
pAcGFP1-1是一種無(wú)啟動(dòng)子載體，可用于監(jiān)測(cè)插入到多克隆位點(diǎn)（MCS）中的不同啟動(dòng)子和啟動(dòng)子/增強(qiáng)子組合的轉(zhuǎn)錄。AcGFP1上游的序列已經(jīng)轉(zhuǎn)化為Kozak共有翻譯起始位點(diǎn)（2），以增強(qiáng)真核細(xì)胞中的翻譯效率。AcGFP1基因下游的SV40多聚腺苷酸信號(hào)直接適當(dāng)處理AcGFP1 mRNA的3'末端。載體主鏈含有用于在表達(dá)SV40T抗原的哺乳動(dòng)物細(xì)胞中復(fù)制的SV40來(lái)源，用于在大腸桿菌中繁殖的pUC起始點(diǎn)和用于單鏈DNA生產(chǎn)的f1起點(diǎn)。新霉素抗性盒（Neor）允許使用G418選擇穩(wěn)定轉(zhuǎn)染的真核細(xì)胞。該盒由SV40早期啟動(dòng)子,Tn5的新霉素/卡那霉素抗性基因和來(lái)自單純皰疹病毒胸苷激酶（HSV TK）基因的多聚腺苷酸化信號(hào)組成。盒上游的細(xì)菌啟動(dòng)子在大腸桿菌中表達(dá)卡那霉素抗性。
pAcGFP1-1 encodes the green fuorescent protein AcGFP1, a derivative of AcGFP from Aequorea coerulescens. AcGFP1 has been optimized for brighter fuorescence. (Excitation maximum = 475 nm; emission maximum = 505 nm.) The coding sequence of the AcGFP1 gene contains silent base changes, which correspond to human codon-usage preferences .
pAcGFP1-1 is a promoterless vector that can be used to monitor transcription from different promoters and promoter/enhancer combinations inserted into the multiple cloning site (MCS). Sequences upstream of AcGFP1 have been converted to a Kozak consensus translation initiation site (2) to enhance translation effciency in eukaryotic cells. SV40 polyadenylation signals downstream of the AcGFP1 gene direct proper processing of the 3' end of the AcGFP1 mRNA. The vector backbone contains an SV40 origin for replication in mammalian cells expressing the SV40 T antigen, a pUC origin of replication for propagation in E. coli, and an f1 origin for single-stranded DNA production. A neomycin-resistance cassette (Neor) allows stably transfected eukaryotic cells to be selected using G418. This cassette consists of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the Herpes simplex virus thymidine kinase (HSV TK) gene. A bacterial promoter upstream of the cassette expresses kanamycin resistance in E. coli.
質(zhì)粒只保證關(guān)鍵序列正確，不保證表達(dá)效果。
■ 質(zhì)粒圖譜
■ 質(zhì)粒序列
質(zhì)粒序列請(qǐng)下載：
ZK421pAcGFP1-1哺乳啟動(dòng)子檢測(cè)質(zhì)粒.txt
