质粒类型：荧光蛋白报告载体

启动子：CMV promoter

克隆方法：多克隆位点，限制性内切酶

载体大小：4733 bp 

5' 测序引物及序列：EGFP-N (5'-CGTCGCCGTCCAGCTCGACCAG-3')

载体标签：ECFP青色荧光 (C端)

载体抗性：Kanamycin (卡那霉素)

筛选标记：Neomycin (新霉素)

pECFP-N1 encodes an enhanced cyan fluorescent variant of the Aequorea victoria green fluorescent protein gene (GFP). The ECFP gene contains six amino acid substitutions. The Tyr-66 to Trp substitution gives ECFP fluorescence excitation (major peak at 433 nm and a minor peak at 453 nm) and emission (major peak at 475 nm and a minor peak at 501 nm) similar to other cyan emission variants (1, 2). The other five substitutions (Phe-64 to Leu; Ser-65 to Thr; Asn-146 to Ile; Met-153 to Thr; and Val-163 to Ala) enhance the brightness and solubility of the protein, primarily due to improved protein folding properties and efficiency of chromophore formation (1, 3, 4). In addition to the chromophore mutations, ECFP contains >190 silent mutations that create an open reading frame comprised almost entirely of preferred human codons (5, 6). Furthermore, upstream sequences flanking ECFP have been converted to a Kozak consensus translation initiation site (7). These changes increase the translational efficiency of the ECFP mRNA and consequently the expression of ECFP in mammalian and plant cells.

The vector contains an SV40 origin of replication and a neomycin resistance (Neor) gene for selection (using G418) in mammalian cells. A bacterial promoter upstream of this cassette (P ) expresses kanamycin resistance in E. coli. The vector backbone also provides a pUC19 origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production. The MCS in pECFP-N1 is between the immediate early promoter of CMV (PCMV IE) and the ECFP coding sequences. Genes cloned into the MCS will be expressed as fusions to the N-terminus of ECFP if they are in the same reading frame as ECFP and there are no intervening stop codons. The inserted gene should include an initiating ATG codon. ECFP fusion proteins retain the fluorescent properties of the native protein in vivo.

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pECFP-N1质粒序列.txt