基本信息
| 启动子: | pAmp |
| 复制子: | pBBR1 Rep，pBBR1 oriV |
| 质粒分类: | pBBR1MCS系列质粒 |
| 质粒大小: | 5996bp |
| 质粒标签: | EGFP |
| 原核抗性: | 卡那霉素Kanamycin |
| 克隆菌株: | DH5α |
| 培养条件: | 37℃，有氧，LB |
| 表达宿主: | 革兰氏阴性菌 |
| 培养条件: | 参考相关文献 |
| 5'测序引物: | M13R:CAGGAAACAGCTATGACC |
| 3'测序引物: | M13F:TGTAAAACGACGGCCAGT |
| 备注: | 低拷贝质粒 |
质粒简介
广宿主穿梭质粒pBBR1MCS2-pAmp-EGFP是基于质粒pBBR1MCS构建的[1]，该质粒是由Kovach等构建，已经证实可以在许多革兰氏阴性菌中进行复制[2]，包括Acetobacter xylinum、Alcaligenes eutrophus、Bartonella bacilliformis、百日咳杆菌（Bordetella spp.）、布鲁氏菌（Brucella spp.）、Caulobacter crescentus、大肠杆菌、Paracoccous denitrificans、荧光假单胞菌（Pseudomonas fluorescens）、恶臭假单胞菌（P. putida）、苜蓿根瘤菌（Rhizobium meliloti）、R. leguminosarum by. Viciae、球形红假单胞菌（Rhodobacter sphaeroides）、鼠伤寒沙门氏菌（Salmonella typhimurium）、霍乱弧菌（Vibrio cholerae）和Xanthomonas campestris等。
[1] Kovach ME, et al. pBBR1MCS: a broad-host-range cloning vector. Biotechniques, 1994,16(5): 800-802.
[2] Kovach ME, Elzer PH, Hill DS, et al. Four new derivatives of the broad-host-range cloning vector pBBR1MCS, carrying different antibiotic-resistance cassettes. Gene, 1995,166:175-176
质粒图谱
质粒序列
LOCUS Exported 5996 bp ds-DNA circular SYN 23-JUL-2016
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pBBR1MCS2-pAmp-EGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5996)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Saturday, September 10, 2016 from SnapGene Viewer 3.1.4
FEATURES Location/Qualifiers
source 1..5996
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 458..1252
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/note="NeoR/KanR"
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
protein_bind 1657..1678
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 1693..1723
/note="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 1731..1747
/bound_moiety="lac repressor encoded by lacI"
/note="lac operator"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 1755..1771
/note="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
promoter 1792..1810
/note="T3 promoter"
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 1850..1954
/gene="bla"
/note="AmpR promoter"
CDS 1955..2671
/codon_start=1
/product="enhanced GFP"
/note="EGFP"
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
misc_feature 2672..2737
/note="MCS"
/note="multiple cloning site"
promoter complement(2791..2809)
/note="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(2819..2835)
/note="M13 fwd"
/note="common sequencing primer, one of multiple similar
variants"
CDS complement(3542..4204)
/codon_start=1
/product="replication protein for the broad-host-range
plasmid pBBR1 from Bordetella bronchiseptica"
/note="pBBR1 Rep"
/translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
rep_origin 4205..4974
/note="pBBR1 oriV"
/note="replication origin of the broad-host-range plasmid
pBBR1 from Bordetella bronchiseptica; requires the pBBR1
Rep protein for replication"
ORIGIN
1 accttcggga gcgcctgaag cccgttctgg acgccctggg gccgttgaat cgggatatgc
61 aggccaaggc cgccgcgatc atcaaggccg tgggcgaaaa gctgctgacg gaacagcggg
121 aagtccagcg ccagaaacag gcccagcgcc agcaggaacg cgggcgcgca catttccccg
181 aaaagtgcca cctgggatga atgtcagcta ctgggctatc tggacaaggg aaaacgcaag
241 cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg cgatagctag actgggcggt
301 tttatggaca gcaagcgaac cggaattgcc agctggggcg ccctctggta aggttgggaa
361 gccctgcaaa gtaaactgga tggctttctt gccgccaagg atctgatggc gcaggggatc
421 aagatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
481 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
541 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
601 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
661 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
721 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
781 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
841 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
901 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
961 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
1021 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
1081 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
1141 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
1201 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
1261 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
1321 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
1381 atcctccagc gcggggatct catgctggag ttcttcgccc acccccatgg gcaaatatta
1441 tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga
1501 tggcttccat gtcggcagaa tgcttaatga attacaacag tttttatgca tgcgcccaat
1561 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
1621 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
1681 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
1741 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
1801 tcactaaagg gaacaaaagc tgggtaccgg gccccccctc gaggtcgacc gcggaacccc
1861 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
1921 ataaatgctt caataatatt gaaaaaggaa gagtatggtg agcaagggcg aggagctgtt
1981 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
2041 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
2101 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt
2161 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
2221 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
2281 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
2341 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
2401 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
2461 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
2521 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag
2581 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
2641 gatcactctc ggcatggacg agctgtacaa gtccggactc agatctcgag ctcaagcttc
2701 gaattctgca gtcgacggta ccgcgggccc gggatccacc ggatctagat aatctagagc
2761 ggccgccacc gcggtggagc tccaattcgc cctatagtga gtcgtattac gcgcgctcac
2821 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
2881 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
2941 cttcccaaca gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta atattttgtt
3001 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgactgcga
3061 tgagtggcag ggcggggcgt aattttttta aggcagttat tggtgccctt aaacgcctgg
3121 tgctacgcct gaataagtga taataagcgg atgaatggca gaaattcgaa agcaaattcg
3181 acccggtcgt cggttcaggg cagggtcgtt aaatagccgc ttatgtctat tgctggttta
3241 ccggtttatt gactaccgga agcagtgtga ccgtgtgctt ctcaaatgcc tgaggccagt
3301 ttgctcaggc tctccccgtg gaggtaataa ttgacgatat gatcatttat tctgcctccc
3361 agagcctgat aaaaacggtg aatccgttag cgaggtgccg ccggcttcca ttcaggtcga
3421 ggtggcccgg ctccatgcac cgcgacgcaa cgcggggagg cagacaaggt atagggcggc
3481 gaggcggcta cagccgatag tctggaacag cgcacttacg ggttgctgcg caacccaagt
3541 gctaccggcg cggcagcgtg acccgtgtcg gcggctccaa cggctcgcca tcgtccagaa
3601 aacacggctc atcgggcatc ggcaggcgct gctgcccgcg ccgttcccat tcctccgttt
3661 cggtcaaggc tggcaggtct ggttccatgc ccggaatgcc gggctggctg ggcggctcct
3721 cgccggggcc ggtcggtagt tgctgctcgc ccggatacag ggtcgggatg cggcgcaggt
3781 cgccatgccc caacagcgat tcgtcctggt cgtcgtgatc aaccaccacg gcggcactga
3841 acaccgacag gcgcaactgg tcgcggggct ggccccacgc cacgcggtca ttgaccacgt
3901 aggccgacac ggtgccgggg ccgttgagct tcacgacgga gatccagcgc tcggccacca
3961 agtccttgac tgcgtattgg accgtccgca aagaacgtcc gatgagcttg gaaagtgtct
4021 tctggctgac caccacggcg ttctggtggc ccatctgcgc cacgaggtga tgcagcagca
4081 ttgccgccgt gggtttcctc gcaataagcc cggcccacgc ctcatgcgct ttgcgttccg
4141 tttgcaccca gtgaccgggc ttgttcttgg cttgaatgcc gatttctctg gactgcgtgg
4201 ccatgcttat ctccatgcgg tagggtgccg cacggttgcg gcaccatgcg caatcagctg
4261 caacttttcg gcagcgcgac aacaattatg cgttgcgtaa aagtggcagt caattacaga
4321 ttttctttaa cctacgcaat gagctattgc ggggggtgcc gcaatgagct gttgcgtacc
4381 cccctttttt aagttgttga tttttaagtc tttcgcattt cgccctatat ctagttcttt
4441 ggtgcccaaa gaagggcacc cctgcggggt tcccccacgc cttcggcgcg gctccccctc
4501 cggcaaaaag tggcccctcc ggggcttgtt gatcgactgc gcggccttcg gccttgccca
4561 aggtggcgct gcccccttgg aacccccgca ctcgccgccg tgaggctcgg ggggcaggcg
4621 ggcgggcttc gccttcgact gcccccactc gcataggctt gggtcgttcc aggcgcgtca
4681 aggccaagcc gctgcgcggt cgctgcgcga gccttgaccc gccttccact tggtgtccaa
4741 ccggcaagcg aagcgcgcag gccgcaggcc ggaggctttt ccccagagaa aattaaaaaa
4801 attgatgggg caaggccgca ggccgcgcag ttggagccgg tgggtatgtg gtcgaaggct
4861 gggtagccgg tgggcaatcc ctgtggtcaa gctcgtgggc aggcgcagcc tgtccatcag
4921 cttgtccagc agggttgtcc acgggccgag cgaagcgagc cagccggtgg ccgctcgcgg
4981 ccatcgtcca catatccacg ggctggcaag ggagcgcagc gaccgcgcag ggcgaagccc
5041 ggagagcaag cccgtagggc gccgcagccg ccgtaggcgg tcacgacttt gcgaagcaaa
5101 gtctagtgag tatactcaag cattgagtgg cccgccggag gcaccgcctt gcgctgcccc
5161 cgtcgagccg gttggacacc aaaagggagg ggcaggcatg gcggcatacg cgatcatgcg
5221 atgcaagaag ctggcgaaaa tgggcaacgt ggcggccagt ctcaagcacg cctaccgcga
5281 gcgcgagacg cccaacgctg acgccagcag gacgccagag aacgagcact gggcggccag
5341 cagcaccgat gaagcgatgg gccgactgcg cgagttgctg ccagagaagc ggcgcaagga
5401 cgctgtgttg gcggtcgagt acgtcatgac ggccagcccg gaatggtgga agtcggccag
5461 ccaagaacag caggcggcgt tcttcgagaa ggcgcacaag tggctggcgg acaagtacgg
5521 ggcggatcgc atcgtgacgg ccagcatcca ccgtgacgaa accagcccgc acatgaccgc
5581 gttcgtggtg ccgctgacgc aggacggcag gctgtcggcc aaggagttca tcggcaacaa
5641 agcgcagatg acccgcgacc agaccacgtt tgcggccgct gtggccgatc tagggctgca
5701 acggggcatc gagggcagca aggcacgtca cacgcgcatt caggcgttct acgaggccct
5761 ggagcggcca ccagtgggcc acgtcaccat cagcccgcaa gcggtcgagc cacgcgccta
5821 tgcaccgcag ggattggccg aaaagctggg aatctcaaag cgcgttgaga cgccggaagc
5881 cgtggccgac cggctgacaa aagcggttcg gcaggggtat gagcctgccc tacaggccgc
5941 cgcaggagcg cgtgagatgc gcaagaaggc cgatcaagcc caagagacgg cccgag
//
温馨提示：不可用于临床ZL。
