| 质粒名称: | pAra-tns |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 170632 |
| 存储实验室: | Sheng Yang |
|
载体骨架基本信息 |
|
| 载体名称: | pCOLADuet-1 |
| 完整质粒大小: | 7748 bp |
| 载体类型: | 细菌表达 |
| 载体抗性: | Kanamycin |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | VchTnsABC |
| 物种来源: | Vibrio cholera |
| 克隆方法: | Gibson Cloning |
|
文献引用方法 |
|
| 材料和方法部分: | pAra-tns was a gift from Sheng Yang (Addgene plasmid # 170632 ; http://n2t.net/addgene:170632 ; RRID:Addgene_170632) |
| 参考文献部分: | Programming Cells by Multicopy Chromosomal Integration Using CRISPR-Associated Transposases. Zhang Y, Yang J, Yang S, Zhang J, Chen J, Tao R, Jiang Y, Yang J, Yang S. CRISPR J. 2021 Jun;4(3):350-359. doi: 10.1089/crispr.2021.0018. 10.1089/crispr.2021.0018 PubMed 34152213 |
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载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pAra-tns 7748 bp ds-DNA circular SYN 03-AUG-2021
DEFINITION Expression of CRISPR-associated transposases.
ACCESSION .
VERSION .
KEYWORDS pAra-tns
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7748)
AUTHORS Zhang Y, Yang J, Yang S, Zhang J, Chen J, Tao R, Jiang Y, Yang J,
Yang S
TITLE Programming Cells by Multicopy Chromosomal Integration Using
CRISPR-Associated Transposases.
JOURNAL CRISPR J. 2021 Jun;4(3):350-359. doi: 10.1089/crispr.2021.0018.
PUBMED 34152213
REFERENCE 2 (bases 1 to 7748)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 3, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7748
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS complement(417..1265)
/codon_start=1
/gene="E. coli rhaR"
/product="transcriptional activator of rhaR and rhaS"
/label=rhaR
/note="needed for rhamnose-dependent activation of the rhaB
promoter"
/translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
D"
CDS complement(1339..2175)
/codon_start=1
/gene="E. coli rhaS"
/product="positive regulator of the rhaB promoter
"
/label=rhaS
/note="activated by L-rhamnose"
/translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
promoter 2321..2439
/label=rhaB promoter
/note="promoter of the E. coli rhaBAD operon, conferring
tight induction with L-rhamnose and repression with
D-glucose in the presence of RhaR and RhaS (Giacalone et
al., 2006)"
CDS 4561..4572
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(5999..6017)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 6013..6060
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS complement(6290..7105)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 7013..7032
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
promoter complement(7106..7197)
/gene="bla"
/label=AmpR promoter
rep_origin complement(7215..102)
/direction=LEFT
/label=ColA ori
/note="Plasmids containing the ColA origin of replication
can be propagated in E. coli cells that contain additional
plasmids with compatible origins."
primer_bind complement(7360..7379)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 cgatcaaacc gcctccccag gcggtttttt cgtttacaga gcaggagatt acgacgatcg
61 taaaaggatc tcaagaagat cctttacgga ttcccgacac catcactcta gatttcagtg
121 caatttatct cttcaaatgt agcacctgaa gtcagcccca tacgatataa gttgtaattc
181 tcatgttagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
241 gcatcggtcg agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgcg
301 ttcttatggc tcttgtatct atcagtgaag catcaagact aacaaacaaa agtagaacaa
361 ctgttcaccg ttacatatca aagggaaaac tgtccatacc catggggcat ggcgaattaa
421 tctttctgcg aattgagatg acgccactgg ctgggcgtca tcccggtttc ccgggtaaac
481 accaccgaaa aatagttact atcttcaaag ccacattcgg tcgaaatatc actgattaac
541 aggcggctat gctggagaag atattgcgca tgacacactc tgacctgtcg cagatattga
601 ttgatggtca ttccagtctg ctggcgaaat tgctgacgca aaacgcgctc actgcacgat
661 gcctcatcac aaaatttatc cagcgcaaag ggacttttca ggctagccgc cagccgggta
721 atcagcttat ccagcaacgt ttcgctggat gttggcggca acgaatcact ggtgtaacga
781 tggcgattca gcaacatcac caactgcccg aacagcaact cagccatttc gttagcaaac
841 ggcacatgct gactactttc atgctcaagc tgaccgataa cctgccgcgc ctgcgccatc
901 cccatgctac ctaagcgcca gtgtggttgc cctgcgctgg cgttaaatcc cggaatcgcc
961 ccctgccagt caagattcag cttcagacgc tccgggcaat aaataatatt ctgcaaaacc
1021 agatcgttaa cggaagcgta ggagtgttta tcgtcagcat gaatgtaaaa gagatcgcca
1081 cgggtaatgc gataagggcg atcgttgagt acatgcaggc cattaccgcg ccagacaatc
1141 accagctcac aaaaatcatg tgtatgttca gcaaagacat cttgcggata acggtcagcc
1201 acagcgactg cctgctggtc gctggcaaaa aaatcatctt tgagaagttt taactgatgc
1261 gccaccgtgg ctacctcggc cagagaacga agttgattat tcgcaatatg gcgtacaaat
1321 acgttgagaa gattcgcgtt attgcagaaa gccatcccgt ccctggcgaa tatcacgcgg
1381 tgaccagtta aactctcggc gaaaaagcgt cgaaaagtgg ttactgtcgc tgaatccaca
1441 gcgataggcg atgtcagtaa cgctggcctc gctgtggcgt agcagatgtc gggctttcat
1501 cagtcgcagg cggttcaggt atcgctgagg cgtcagtccc gtttgctgct taagctgccg
1561 atgtagcgta cgcagtgaaa gagaaaattg atccgccacg gcatcccaat tcacctcatc
1621 ggcaaaatgg tcctccagcc aggccagaag caagttgaga cgtgatgcgc tgttttccag
1681 gttctcctgc aaactgcttt tacgcagcaa gagcagtaat tgcataaaca agatctcgcg
1741 actggcggtc gagggtaaat cattttcccc ttcctgctgt tccatctgtg caaccagctg
1801 tcgcacctgc tgcaatacgc tgtggttaac gcgccagtga gacggatact gcccatccag
1861 ctcttgtggc agcaactgat tcagcccggc gagaaactga aatcgatccg gcgagcgata
1921 cagcacattg gtcagacaca gattatcggt atgttcatac agatgccgat catgatcgcg
1981 tacgaaacag accgtgccac cggtgatggt atagggctgc ccattaaaca catgaatacc
2041 cgtgccatgt tcgacaatca caatttcatg aaaatcatga tgatgttcag gaaaatccgc
2101 ctgcgggagc cggggttcta tcgccacgga cgcgttacca gacggaaaaa aatccacact
2161 atgtaatacg gtcatactgg cctcctgatg tcgtcaacac ggcgaaatag taatcacgag
2221 gtcaggttct taccttaaat tttcgacgga aaaccacgta aaaaacgtcg atttttcaag
2281 atacagcgtg aattttcagg aaatgcggtg agcatcacat caccacaatt cagcaaattg
2341 tgaacatcat cacgttcatc tttccctggt tgccaatggc ccattttcct gtcagtaacg
2401 agaaggtcgc gaattcaggc gctttttaga ctggtcgtac ctgtagaaat aattttgttt
2461 aactttaata aggagatata ccatggcgac aagtttacct acgccctcag caattacgac
2521 ttcggcgtta gagtatgcat tccatactcc cgctcgcaat ctaacgaaat ctcgcggaaa
2581 aaacattcat cgttatgtca gtgtaaagat gagtaagagg attacggtag aatctactct
2641 agagtgtgat gcctgctatc actttgattt tgagccaagt attgttcgct tttgcgctca
2701 accgattcga tttttatatt atctcaatgg tcagtctcac tcctatgttc ctgactttct
2761 agttcaattt gataccaacg agtttgttct atatgaagta aagtcagctt atgctaagaa
2821 caaacctgat tttgatgttg aatgggaggc gaaagtaaaa gcagcaactg aactagggct
2881 agaattggag cttgttgaag agagtgatat tagggatacg gttgtattaa ataatcttaa
2941 gcgcatgcat cgttatgctt cgaaagatga gctgaataac gtacataact ctctcttaaa
3001 aataataaag tacaatggcg cccaatctgc aagatgcttg ggagaacagt tgggtttaaa
3061 aggccgaact gttttaccaa ttttgtgcga tttgctgtca aggtgtttac tcgatacacg
3121 tttggataag cctctatctc ttgaatctcg atttgagttg gccagttatg gctaagaaag
3181 ggttctcaag tttccataga aaagcagtct cttcccaaga tacgcttgaa tcgatagagc
3241 ttgtctctag cgctaattgt ctagaaagtg ttacgtatca agatatatca gcatttcccg
3301 aaacaattgc ggtagagatt aatttccgat taagcattct tcgcttttta gcgcgaaagt
3361 gcgaaaccat tgtggccaaa tcaattgaac cacatcgtgt agagctacag caaaactata
3421 gtagaaaaat acccagtgca ataacgatat atcgatggtg gcttgctttt cgaaaatcag
3481 actacaaccc cattagctta gcacctaata tcaaggatag aggtaataga gagacaaaag
3541 tgtcaacagt tgttgattct attatggaac aggcagttga aagagttata tctggacgaa
3601 aagtcaatgt tagctctgca tataaacgtg ttcgacgaaa agttcgtcaa tacaatctaa
3661 ctcatggaac gaaatacacg tatcctaagt acgaatctgt aagaaagcga gtaaaaaaga
3721 aaaccccatt tgagttatta gccgcaggga aaggggagag agtagctaag agagagtttc
3781 gccgaatggg aaaaaagatc ctcacgtcta gcgtgctaga gagggttgaa atagatcaca
3841 ctgtcgttga cctttttgca gtacacgaag agtatcgaat cccattgggc cgaccttggc
3901 ttactcaatt ggttgattgt tacagtaaag ctgtaatcgg tttttattta ggtttcgagc
3961 ctcctagcta tgtgtcggtt tcccttgcac ttaagaatgc aatacaacgc aaagatgact
4021 taatctcctc gtatgaatcg atcgagaatg aatggctatg ttatggcatc ccagacctac
4081 tcgtaactga taatggtaaa gagtttttgt cgaaagcatt tgatcaagca tgtgaatcac
4141 tattgatcaa tgtgcatcaa aataaagttg agacgcccga caacaaacct catgttgaac
4201 gtaactacgg gactattaat acttctctgt tagacgattt acctgggaaa tccttcagcc
4261 agtaccttca aagagaaggg tacgactctg tgggagaagc tacccttaca ctcaatgaga
4321 ttagagaaat ttacttaatt tggttggtgg atatttatca taaaaaaccc aatcagagag
4381 gcactaattg tcctaatgtt gcgtggaaaa agggttgtca agaatgggaa ccagaggagt
4441 tctctggttc taaagacgaa ttagacttta aatttgctat tgttgattac aaacaactta
4501 ctaaagtagg gataactgtc tacaaagaac tgagttatag caatgaccgt ttagctgaat
4561 atagagggaa gaaaggaaac cataaagttc agttcaagta taaccctgag tgtatggcag
4621 ttatttgggt gttggatgag gatatgaatg agtactttac agttaatgcg attgactacg
4681 aatatgcaag tagagtatca ctttggcaac ataaatataa catgaaatat caagcagaac
4741 taaattcagc agaatatgat gaggacaagg aaattgatgc agaaataaaa attgaagaaa
4801 tcgcagatcg ttcaattgtt aagactaaca aaatcagagc tcggaggcgt ggcgctaggc
4861 atcaagagaa tagcgcaagg gctaagtcaa tcagtaatgc gaacccggcc tcgatacaaa
4921 aacatgaaga tgaaatcgtt agtgcagata atgacgattg ggatattgat tatgtctgag
4981 aaatcgtcaa atgagtgaaa cgcgtgaggc tcgtatatca agagctaaaa gggcatttgt
5041 atccacaccg agcgttagga aaatcttgag ttacatggat agatgtagag atctatcaga
5101 cctagagtct gagcctacat gcatgatggt ctatggtgct tcgggtgtag gtaaaacgac
5161 cgtcatcaag aaatacttaa atcaaaacag aagagagtcc gaagccgggg gcgatataat
5221 accggttttg catattgagt tgccagacaa tgcgaagcca gtagatgcag caagggaatt
5281 gctggttgaa atgggtgacc cgctagcact ttatgaaact gacttagcta gattgacgaa
5341 aagactgact gaattaatcc ctgcggtcgg cgtgaagctg attattatcg atgagttcca
5401 acatttggtg gaagaaaggt caaatcgggt tcttacccaa gtaggtaatt ggctaaaaat
5461 gatacttaac aaaacgaaat gtccaattgt tatatttggt atgccatact caaaagttgt
5521 actgcaagca aactcgcaac ttcacgggcg attttccatt caggttgaac tgcgcccctt
5581 tagctaccag ggaggtagag gtgtatttaa aacttttttg gaataccttg ataaagccct
5641 accttttgaa aaacaggctg gcttagccaa cgaaagtttg cagaaaaaat tgtatgcatt
5701 ctctcaggga aacatgcgtt cgttgagaaa ccttatttat caagcatcta tcgaagcaat
5761 tgataatcag catgagacga taaccgaaga agatttcgtt tttgcatcga agttgacatc
5821 gggcgataaa cccaactcat ggaaaaatcc ttttgaggag ggtgttgagg taacagaaga
5881 tatgttacga ccgccaccaa aagatattgg ttgggaggac tatttgagac attcaacccc
5941 gagagtgagt aaaccaggta gaaataaaaa ctttttcgaa taacctaggc tgctgccacc
6001 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
6061 ctgaaacctc aggcatttga gaagcacacg gtcacactgc ttccggtagt caataaaccg
6121 gtaaaccagc aatagacata agcggctatt taacgaccct gccctgaacc gacgacaagc
6181 tgacgaccgg gtctccgcaa gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
6241 tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact
6301 catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt
6361 gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa
6421 gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc
6481 cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg
6541 agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct
6601 cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga
6661 gacgaaatac gcggtcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc
6721 gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata
6781 cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac
6841 ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca
6901 tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg
6961 catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag
7021 cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag
7081 acgtttcccg ttgaatatgg ctcatactct tcctttttca atattattga agcatttatc
7141 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
7201 gcatgctagc gcagaaacgt cctagaagat gccaggagga tacttagcag agagacaata
7261 aggccggagc gaagccgttt ttccataggc tccgcccccc tgacgaacat cacgaaatct
7321 gacgctcaaa tcagtggtgg cgaaacccga caggactata aagataccag gcgtttcccc
7381 ctgatggctc cctcttgcgc tctcctgttc ccgtcctgcg gcgtccgtgt tgtggtggag
7441 gctttaccca aatcaccacg tcccgttccg tgtagacagt tcgctccaag ctgggctgtg
7501 tgcaagaacc ccccgttcag cccgactgct gcgccttatc cggtaactat catcttgagt
7561 ccaacccgga aagacacgac aaaacgccac tggcagcagc cattggtaac tgagaattag
7621 tggatttaga tatcgagagt cttgaagtgg tggcctaaca gaggctacac tgaaaggaca
7681 gtatttggta tctgcgctcc actaaagcca gttaccaggt taagcagttc cccaactgac
7741 ttaacctt
//
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