质粒简介：

 pEGFP-C3已经优化了编码野生型GFP的红移变体以具有更亮的荧光，在哺乳动物细胞中表达较高（最大激发波长= 488nm，最大释放波长= 507nm）。EGFP基因编码序列含有190多个沉默碱基变化，符合人类密码子使用偏好。EGFP序列的一侧为KozaK，进一步提高了真核细胞中翻译的效率。

pEGFP-C3 encodes a red-shifted variant of wild-type GFP which has been optimized for brighter fluorescence and higher expression in mammalian cells. (Excitation maximum = 488 nm; emission maximum = 507 nm.) pEGFP-C3 encodes the GFPmut1 variant  which contains the double-amino-acid 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. Sequences flanking EGFP have been converted to a Kozak consensus translation initiation site (6) to further increase the translation efficiency in eukaryotic cells. The MCS in pEGFP-C3 is between the EGFP coding sequences and the SV40 poly A. Genes cloned into the MCS will be expressed as fusions to the C terminus of EGFP if they are in the same reading frame as EGFP and there are no intervening stop codons. 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), consisting 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, allows stably transfected eukaryotic cells to be selected using G418. A bacterial promoter upstream of this cassette expresses kanamycin resistance in E. coli. The pEGFP-C3 backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.

Fusions to the C terminus of EGFP retain the fluorescent properties of the native protein allowing the localization of the fusion protein in vivo. The target gene should be cloned into pEGFP-C3 so that it is in frame with the EGFP coding sequences, with no intervening in-frame stop codons. The recombinant EGFP vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418. pEGFP-C3 can also be used simply to express EGFP in a cell line of interest (e.g., as a transfection marker).

Propagation in E. coli:Suitable host strains: DH5α, HB101, and other general purpose strains. Single-stranded DNA production requires a host containing an F plasmid such as JM109 or XL1-Blue. Selectable marker: plasmid confers resistance to kanamycin (30 µg/ml) to E. coli hosts.

产品参数：

出品公司：Clontech

启动子: CMV promoter

复制子: pUC ori,f1 ori

终止子: SV40 poly(A) signal

质粒分类: 哺乳系列质粒；哺乳荧光质粒；哺乳绿色质粒

质粒大小: 4727bp

质粒标签: C-EGFP

原核抗性: 卡纳霉素Kan（50μg/ml）

筛选标记: 新霉素Neo/G418

克隆菌株: DH5α等大肠杆菌

培养条件: 37℃，有氧 LB

表达宿主: 293T等哺乳细胞

诱导方式: 无须诱导，瞬时表达

5'测序引物: pEGFP-C-5'（CATGGTCCTGCTGGAGTTCGTG）

3'测序引物: pEGFP-C-3' (TATGGCTGATTATGATCAGT)