■ 基本信息
復制子: | pUC |
終止子: | SV40 poly(A) signal |
質粒分類: | 哺乳系列質粒；哺乳熒光質粒；哺乳綠色質粒 |
質粒大小: | 4151bp |
質粒標簽: | C-EGFP |
原核抗性: | Kan |
真核抗性: | G418 |
克隆菌株: | DH5a |
培養(yǎng)條件: | 37℃，5%CO2 |
表達宿主: | 293T等哺乳細胞 |
誘導方式: | 無須誘導，瞬時表達 |
5'測序引物:
| EGFP-N（CGTCGCCGTCCAGCTCGACCAG）
|
3'測序引物: | EGFP-F (CCAGCAACGCGGCCTTTTTA) |
■ 質粒屬性
質粒宿主: | 哺乳細胞 |
質粒用途: | 信號報告 |
片段類型: | Promoter
|
片段物種: | 空載體
|
原核抗性: | Kan |
真核抗性: | G418 |
熒光標記: |
|
■ 質粒簡介
pEGFP-1編碼野生型GFP（1-3）的紅移型變體，其已經(jīng)針對更明亮的熒光和哺乳動物細胞中更高的表達進行了優(yōu)化。（激發(fā)最大值= 488nm;發(fā)射最大值= 507nm）pEGFP-1骨架還提供了用于在大腸桿菌中繁殖的pUC起始點和用于單鏈DNA生產(chǎn)的f1起源。
pEGFP-1 encodes a red-shifted variant of wild-type GFP (1–3) which has been optimized for brighter fluorescence and higher expression in mammalian cells. (Excitation maximum = 488 nm; emission maximum = 507 nm.) pEGFP-1 encodes the GFPmut1 variant (4) which contains the double-aminoacid substitution of Phe-64 to Leu and Ser-65 to Thr. The coding sequence of the EGFP gene contains more than 190 silent base changes which correspond to human codon-usage preferences (5). Sequences flanking EGFP have been converted to a Kozak consensus translation initiation site (6) to further increase the translation efficiency in eukaryotic cells. pEGFP-1 is a promoterless EGFP vector which can be used to monitor transcription from different promoters and promoter/enhancer combinations inserted into the MCS located upstream of the EGFP coding sequence. SV40 polyadenylation signals downstream of the EGFP gene direct proper processing of the 3' end of the EGFP mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T antigen. A neomycin-resistance cassette (Neor) allows stably transfected eukaryotic cells to be selected using G418. The Neor 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 this cassette confers kanamycin resistance in E. coli. The pEGFP-1 backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.
EGFP can be used as an in vivo reporter of gene expression . Promoters should be cloned into the pEGFP-1 MCS upstream from the EGFP coding sequences. Without the addition of a functional promoter, this vector will not express EGFP. The recombinant EGFP vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418.
質粒只保證關鍵序列正確，不保證表達效果。
■ 質粒圖譜
■ 質粒序列
質粒序列請下載：
ZK410pEGFP-1哺乳啟動子檢測質粒.txt
