质粒类型：逆转录病毒载体

高拷贝/低拷贝：高拷贝

克隆方法：多克隆位点，限制性内切酶

载体大小：5934bp

3' 测序引物及序列：IRES-R: CCTCACATTGCCAAAAGACG

载体抗性：Ampicillin (氨苄青霉素)

备注：载体表达DsRed荧光蛋白作为筛选标志，用来构建稳表达细胞系

pRetroX-IRES-DsRedExpress is a bicistronic, fl uorescent, retroviral vector that allows both a gene of interest and the DsRed-Express gene to be translated from a single bicistronic mRNA. pRetroX-IRES-DsRedExpress is designed for effi cient delivery and selection (by fl ow cytometry or other methods) of stably transduced mammalian cells expressing the DsRed-Express fl uorescent protein and the protein of interest. This vector can be used to obtain stable cell lines without time-consuming drug and clonal selection. DsRed-Express is a rapidly maturing variant of Discosoma sp. red fl uorescent protein (DsRed) and is easily detected with standard Rhodamine/propidium iodide fi lter sets (1).

Bicistronic expression from this vector is facilitated by the encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES). This IRES facilitates cap-independent translation from an internal start site at the IRES/DsRed-Express junction (2). DsRed-Express contains nine amino acid substitutions that enhance its solubility, reduce its green emission, and accelerate its maturation (3). This retroviral vector is derived from the pMIN series of vectors (4, 5). These optimized vectors have the ability to produce high viral titers, express genes at high levels, and, due to the absence of retroviral coding sequences, exhibit improved safety profi les. The multiple cloning site (MCS) in pRetroX-IRES-DsRedExpress is between the 5’ MMLV LTR and the IRES sequence. Genes cloned into the MCS are expressed as a bicistronic message transcribed from the 5’ LTR.

pRetroX-IRES-DsRedExpress contains all of the necessary viral RNA processing elements; these include the 5’ and 3’ LTRs, the packaging signal (ψ), and the tRNA primer-binding site. For safety reasons, however, the vector lacks the structural genes (gag, pol, and env) necessary for retroviral particle formation and replication. pRetroX-IRES-DsRedExpress contains a ColE1 origin of replication, and an E. coli Ampr gene for propagation and selection in bacteria.

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pRetroX-IRES-DsRed Express质粒序列.txt