pZsGreen1-1
pZsGreen1-1
编号 | 载体名称 |
北京华越洋VECT75376 | pZsGreen1-1 |
pZsGreen1-1载体基本信息:
载体名称: | pZsGreen1-1 |
质粒类型: | 无启动子载体;荧光报告载体 |
高拷贝/低拷贝: | 高拷贝 |
克隆方法: | 限制性内切酶,多克隆位点 |
启动子: | 无启动子 |
载体大小: | 4126 bp |
5' 测序引物及序列: | -- |
3' 测序引物及序列: | -- |
载体标签: | 绿色荧光蛋白ZsGreen1 |
载体抗性: | 卡那霉素 |
筛选标记: | Neomycin_1.html' target='_blank'>新霉素(Neomycin) |
克隆菌株: | DH5α, JM109 |
宿主细胞(系): | 真核细胞 |
备注: | pZsGreen1-1载体不含启动子,用于研究目的启动子及顺式转录元件在细胞内的功能和水平; |
稳定性: | 瞬表达 或 稳表达 |
组成型/诱导型: | -- |
病毒/非病毒: | 非病毒 |
pZsGreen1-1载体质粒图谱和多克隆位点信息
pZsGreen1-1载体简介:
pZsGreen1-1 encodes a human codon-optimized variant of wild-type Zoanthus sp. green fluorescent
protein, ZsGreen1 (1). The ZsGreen1 coding sequence contains a series of silent base-pair changes,
which correspond to human codon-usage preferences, for optimal expression in mammalian cells
(2). A single amino acid substitution (Asn-66 to Met) has been made to enhance the emission
characteristics of ZsGreen1 (excitation maximum = 493 nm; emission maximum = 505 nm).
pZsGreen1-1 is a promoterless ZsGreen1 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 ZsGreen1 have been converted to a Kozak consensus translation
initiation site (3) to increase translation efficiency in eukaryotic cells. SV40 polyadenylation signals
downstream of the ZsGreen1 gene direct proper processing of the 3' end of the ZsGreen1 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 singlestranded
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.
pZsGreen1-1载体使用:
ZsGreen1 can be used as an in vivo reporter of gene expression. Promoters should be cloned
into the pZsGreen1-1 MCS upstream from the ZsGreen1 coding sequence. Without addition of a
functional promoter, this vector will not express ZsGreen1. The recombinant ZsGreen1 vector can
be transfected into mammalian cells using any standard transfection method. If required, stable
transfectants can be selected using G418 (4).
pZsGreen1-1载体特征:
MCS: 12–83
Zoanthus sp. human codon-optimized Green Fluorescent Protein (ZsGreen1) gene
Kozak consensus translation initiation site: 90–100
Start codon (ATG): 97–99; Stop codon: 790–792
Asn-66 to Met mutation (A→T, C→G ): 293, 294
SV40 early mRNA polyadenylation signal
Polyadenylation signals: 945–950 & 974–979
mRNA 3' ends: 983 & 995
f1 single-strand DNA origin: 1042–1497
(Packages noncoding strand of ZsGreen1.)
Ampicillin resistance (β-lactamase) promoter
–35 region: 1559–1564; –10 region: 1582–1587
Transcription start point: 1594
SV40 origin of replication: 1838–1973
SV40 early promoter
Enhancer (72-bp tandem repeats): 1669–1742 & 1743–1814
21-bp repeats: 1818–1837, 1839–1859 & 1861–1881
Early promoter element: 1894–1900
Major transcription start points: 1890, 1928, 1934 & 1939
Kanamycin/neomycin resistance geneNeomycin phosphotransferase coding sequences:
Start codon (ATG): 2022–2024; stop codon: 2814–2816
G→A mutation to remove Pst I site: 2204
C→A (Arg→Ser) mutation to remove BssH II site: 2550
Herpes simplex virus (HSV) thymidine kinase (TK) polyadenylation signal
Polyadenylation signals: 3052–3057 & 3065–3070
pUC plasmid replication origin: 3401–4044
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 (50 μg/ml) in E. coli hosts.
E. coli replication origin: pUC
Copy number: ≈500
Plasmid incompatibility group: pMB1/Col E1
Green Fluorescent Protein (ZsGreen1)
Excitation/Emission Maxima: 493 nm / 505 nm
pZsGreen1-1载体序列:
ORIGIN
1 TAGTTATTAC TAGCGCTACC GGACTCAGAT CTCGAGCTCA AGCTTCGAAT TCTGCAGTCG
61 ACGGTACCGC GGGCCCGGGA TCCACCGGTC GCCACCATGG CCCAGTCCAA GCACGGCCTG
121 ACCAAGGAGA TGACCATGAA GTACCGCATG GAGGGCTGCG TGGACGGCCA CAAGTTCGTG
181 ATCACCGGCG AGGGCATCGG CTACCCCTTC AAGGGCAAGC AGGCCATCAA CCTGTGCGTG
241 GTGGAGGGCG GCCCCTTGCC CTTCGCCGAG GACATCTTGT CCGCCGCCTT CATGTACGGC
301 AACCGCGTGT TCACCGAGTA CCCCCAGGAC ATCGTCGACT ACTTCAAGAA CTCCTGCCCC
361 GCCGGCTACA CCTGGGACCG CTCCTTCCTG TTCGAGGACG GCGCCGTGTG CATCTGCAAC
421 GCCGACATCA CCGTGAGCGT GGAGGAGAAC TGCATGTACC ACGAGTCCAA GTTCTACGGC
481 GTGAACTTCC CCGCCGACGG CCCCGTGATG AAGAAGATGA CCGACAACTG GGAGCCCTCC
541 TGCGAGAAGA TCATCCCCGT GCCCAAGCAG GGCATCTTGA AGGGCGACGT GAGCATGTAC
601 CTGCTGCTGA AGGACGGTGG CCGCTTGCGC TGCCAGTTCG ACACCGTGTA CAAGGCCAAG
661 TCCGTGCCCC GCAAGATGCC CGACTGGCAC TTCATCCAGC ACAAGCTGAC CCGCGAGGAC
721 CGCAGCGACG CCAAGAACCA GAAGTGGCAC CTGACCGAGC ACGCCATCGC CTCCGGCTCC
781 GCCTTGCCCT GAGCGGCCGC GACTCTAGAT CATAATCAGC CATACCACAT TTGTAGAGGT
841 TTTACTTGCT TTAAAAAACC TCCCACACCT CCCCCTGAAC CTGAAACATA AAATGAATGC
901 AATTGTTGTT GTTAACTTGT TTATTGCAGC TTATAATGGT TACAAATAAA GCAATAGCAT
961 CACAAATTTC ACAAATAAAG CATTTTTTTC ACTGCATTCT AGTTGTGGTT TGTCCAAACT
1021 CATCAATGTA TCTTAAGGCG TAAATTGTAA GCGTTAATAT TTTGTTAAAA TTCGCGTTAA
1081 ATTTTTGTTA AATCAGCTCA TTTTTTAACC AATAGGCCGA AATCGGCAAA ATCCCTTATA
1141 AATCAAAAGA ATAGACCGAG ATAGGGTTGA GTGTTGTTCC AGTTTGGAAC AAGAGTCCAC
1201 TATTAAAGAA CGTGGACTCC AACGTCAAAG GGCGAAAAAC CGTCTATCAG GGCGATGGCC
1261 CACTACGTGA ACCATCACCC TAATCAAGTT TTTTGGGGTC GAGGTGCCGT AAAGCACTAA
1321 ATCGGAACCC TAAAGGGAGC CCCCGATTTA GAGCTTGACG GGGAAAGCCG GCGAACGTGG
1381 CGAGAAAGGA AGGGAAGAAA GCGAAAGGAG CGGGCGCTAG GGCGCTGGCA AGTGTAGCGG
1441 TCACGCTGCG CGTAACCACC ACACCCGCCG CGCTTAATGC GCCGCTACAG GGCGCGTCAG
1501 GTGGCACTTT TCGGGGAAAT GTGCGCGGAA CCCCTATTTG TTTATTTTTC TAAATACATT
1561 CAAATATGTA TCCGCTCATG AGACAATAAC CCTGATAAAT GCTTCAATAA TATTGAAAAA
1621 GGAAGAGTCC TGAGGCGGAA AGAACCAGCT GTGGAATGTG TGTCAGTTAG GGTGTGGAAA
1681 GTCCCCAGGC TCCCCAGCAG GCAGAAGTAT GCAAAGCATG CATCTCAATT AGTCAGCAAC
1741 CAGGTGTGGA AAGTCCCCAG GCTCCCCAGC AGGCAGAAGT ATGCAAAGCA TGCATCTCAA
1801 TTAGTCAGCA ACCATAGTCC CGCCCCTAAC TCCGCCCATC CCGCCCCTAA CTCCGCCCAG
1861 TTCCGCCCAT TCTCCGCCCC ATGGCTGACT AATTTTTTTT ATTTATGCAG AGGCCGAGGC
1921 CGCCTCGGCC TCTGAGCTAT TCCAGAAGTA GTGAGGAGGC TTTTTTGGAG GCCTAGGCTT
1981 TTGCAAAGAT CGATCAAGAG ACAGGATGAG GATCGTTTCG CATGATTGAA CAAGATGGAT
2041 TGCACGCAGG TTCTCCGGCC GCTTGGGTGG AGAGGCTATT CGGCTATGAC TGGGCACAAC
2101 AGACAATCGG CTGCTCTGAT GCCGCCGTGT TCCGGCTGTC AGCGCAGGGG CGCCCGGTTC
2161 TTTTTGTCAA GACCGACCTG TCCGGTGCCC TGAATGAACT GCAAGACGAG GCAGCGCGGC
2221 TATCGTGGCT GGCCACGACG GGCGTTCCTT GCGCAGCTGT GCTCGACGTT GTCACTGAAG
2281 CGGGAAGGGA CTGGCTGCTA TTGGGCGAAG TGCCGGGGCA GGATCTCCTG TCATCTCACC
2341 TTGCTCCTGC CGAGAAAGTA TCCATCATGG CTGATGCAAT GCGGCGGCTG CATACGCTTG
2401 ATCCGGCTAC CTGCCCATTC GACCACCAAG CGAAACATCG CATCGAGCGA GCACGTACTC
2461 GGATGGAAGC CGGTCTTGTC GATCAGGATG ATCTGGACGA AGAGCATCAG GGGCTCGCGC
2521 CAGCCGAACT GTTCGCCAGG CTCAAGGCGA GCATGCCCGA CGGCGAGGAT CTCGTCGTGA
2581 CCCATGGCGA TGCCTGCTTG CCGAATATCA TGGTGGAAAA TGGCCGCTTT TCTGGATTCA
2641 TCGACTGTGG CCGGCTGGGT GTGGCGGACC GCTATCAGGA CATAGCGTTG GCTACCCGTG
2701 ATATTGCTGA AGAGCTTGGC GGCGAATGGG CTGACCGCTT CCTCGTGCTT TACGGTATCG
2761 CCGCTCCCGA TTCGCAGCGC ATCGCCTTCT ATCGCCTTCT TGACGAGTTC TTCTGAGCGG
2821 GACTCTGGGG TTCGAAATGA CCGACCAAGC GACGCCCAAC CTGCCATCAC GAGATTTCGA
2881 TTCCACCGCC GCCTTCTATG AAAGGTTGGG CTTCGGAATC GTTTTCCGGG ACGCCGGCTG
2941 GATGATCCTC CAGCGCGGGG ATCTCATGCT GGAGTTCTTC GCCCACCCTA GGGGGAGGCT
3001 AACTGAAACA CGGAAGGAGA CAATACCGGA AGGAACCCGC GCTATGACGG CAATAAAAAG
3061 ACAGAATAAA ACGCACGGTG TTGGGTCGTT TGTTCATAAA CGCGGGGTTC GGTCCCAGGG
3121 CTGGCACTCT GTCGATACCC CACCGAGACC CCATTGGGGC CAATACGCCC GCGTTTCTTC
3181 CTTTTCCCCA CCCCACCCCC CAAGTTCGGG TGAAGGCCCA GGGCTCGCAG CCAACGTCGG
3241 GGCGGCAGGC CCTGCCATAG CCTCAGGTTA CTCATATATA CTTTAGATTG ATTTAAAACT
3301 TCATTTTTAA TTTAAAAGGA TCTAGGTGAA GATCCTTTTT GATAATCTCA TGACCAAAAT
3361 CCCTTAACGT GAGTTTTCGT TCCACTGAGC GTCAGACCCC GTAGAAAAGA TCAAAGGATC
3421 TTCTTGAGAT CCTTTTTTTC TGCGCGTAAT CTGCTGCTTG CAAACAAAAA AACCACCGCT
3481 ACCAGCGGTG GTTTGTTTGC CGGATCAAGA GCTACCAACT CTTTTTCCGA AGGTAACTGG
3541 CTTCAGCAGA GCGCAGATAC CAAATACTGT TCTTCTAGTG TAGCCGTAGT TAGGCCACCA
3601 CTTCAAGAAC TCTGTAGCAC CGCCTACATA CCTCGCTCTG CTAATCCTGT TACCAGTGGC
3661 TGCTGCCAGT GGCGATAAGT CGTGTCTTAC CGGGTTGGAC TCAAGACGAT AGTTACCGGA
3721 TAAGGCGCAG CGGTCGGGCT GAACGGGGGG TTCGTGCACA CAGCCCAGCT TGGAGCGAAC
3781 GACCTACACC GAACTGAGAT ACCTACAGCG TGAGCTATGA GAAAGCGCCA CGCTTCCCGA
3841 AGGGAGAAAG GCGGACAGGT ATCCGGTAAG CGGCAGGGTC GGAACAGGAG AGCGCACGAG
3901 GGAGCTTCCA GGGGGAAACG CCTGGTATCT TTATAGTCCT GTCGGGTTTC GCCACCTCTG
3961 ACTTGAGCGT CGATTTTTGT GATGCTCGTC AGGGGGGCGG AGCCTATGGA AAAACGCCAG
4021 CAACGCGGCC TTTTTACGGT TCCTGGCCTT TTGCTGGCCT TTTGCTCACA TGTTCTTTCC
4081 TGCGTTATCC CCTGATTCTG TGGATAACCG TATTACCGCC ATGCAT
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