| 质粒名称: | pOSIP-TH (TetR, HK022) |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 45988 |
| 存储实验室: | Drew Endy, Keith Shearwin |
|
载体骨架基本信息 |
|
| 载体名称: | pOSIP |
| 载体出品公司: | Clontech |
| 空载体大小: | 6925 bp |
| 完整质粒大小: | 6751bp |
| 载体类型: | Bacterial Expression, Synthetic Biology |
| 载体抗性: | Tetracycline, 10 μg/mL |
| 生长温度: | 30 ℃ |
| 宿主菌: | DB3.1 |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 5’测序引物 | TAAACTGCCAGGAATTGGGGATC |
| 3’测序引物 | GAAACGCAAAAAGGCCATCCGTC |
|
文献引用方法 |
|
| 材料和方法部分: | |
| 参考文献部分: | |
质粒图谱.png)
LOCUS pOSIP-TH (TetR, HK022) 6751 bp DNA circular SYN 14-APR-2025
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6751)
AUTHORS St-Pierre F, Cui L, Priest DG, Endy D, Dodd IB, Shearwin KE
TITLE One-step cloning and chromosomal integration of DNA.
JOURNAL ACS Synth Biol. 2013 Sep 20;2(9):537-41. doi: 10.1021/sb400021j.
Epub 2013 May 20.
PUBMED 24050148
REFERENCE 2 (bases 1 to 6751)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 14, 2025 from SnapGene Server 8.0.1
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; doi: "10.1021/sb400021j";
journalName: "ACS Synth Biol"; date: "2013-09-20- 20"; volume: "2";
issue: "9"; pages: "537-41"
COMMENT Sequence Label: pOSIP-TH (TetR, HK022)
FEATURES Location/Qualifiers
source 1..6751
/mol_type="other DNA"
/organism="synthetic DNA construct"
terminator complement(83..169)
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
primer_bind complement(210..228)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind 379..401
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
terminator 492..738
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
protein_bind 773..999
/gene="attP"
/label=HK022 attP
/bound_moiety="HK022 integrase"
/note="attachment site of phage HK022"
protein_bind complement(1029..1076)
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
rep_origin 1087..1475
/label=R6K gamma ori
/note="gamma replication origin from E. coli plasmid R6K;
requires the R6K initiator protein pi for replication"
terminator complement(1532..1626)
/label=lambda t0 terminator
/note="transcription terminator from phage lambda"
CDS complement(1685..2890)
/codon_start=1
/gene="tetA from transposon Tn10"
/product="tetracycline efflux protein"
/label=TcR
/note="confers resistance to tetracycline"
/translation="MNSSTKIALVITLLDAMGIGLIMPVLPTLLREFIASEDIANHFGV
LLALYALMQVIFAPWLGKMSDRFGRRPVLLLSLIGASLDYLLLAFSSALWMLYLGRLLS
GITGATGAVAASVIADTTSASQRVKWFGWLGASFGLGLIAGPIIGGFAGEISPHSPFFI
AALLNIVTFLVVMFWFRETKNTRDNTDTEVGVETQSNSVYITLFKTMPILLIIYFSAQL
IGQIPATVWVLFTENRFGWNSMMVGFSLAGLGLLHSVFQAFVAGRIATKWGEKTAVLLG
FIADSSAFAFLAFISEGWLVFPVLILLAGGGIALPALQGVMSIQTKSHQQGALQGLLVS
LTNATGVIGPLLFAVIYNHSLPIWDGWIWIIGLAFYCIIILLSMTFMLTPQAQGSKQET
SA"
promoter 2898..2952
/gene="tetR"
/label=tetR/tetA promoters
/note="overlapping promoters for bacterial tetR and tetA"
protein_bind 2899..2917
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
CDS complement(3035..4108)
/codon_start=1
/gene="int"
/product="integrase from phage HK022"
/label=HK022 integrase
/translation="MGRRRSHERRDLPPNLYIRNNGYYCYRDPRTGKEFGLGRDRRIAI
TEAIQANIELLSGNRRESLIDRIKGADAITLHAWLDRYETILSERGIRPKTLLDYASKI
RAIRRKLPDKPLADISTKEVAAMLNTYVAEGKSASAKLIRSTLVDVFREAIAEGHVATN
PVTATRTAKSEVRRSRLTANEYVAIYHAAEPLPIWLRLAMDLAVVTGQRVGDLCRMKWS
DINDNHLHIEQSKTGAKLAIPLTLTIDALNISLADTLQQCREASSSETIIASKHHDPLS
PKTVSKYFTKARNASGLSFDGNPPTFHELRSLSARLYRNQIGDKFAQRLLGHKSDSMAA
RYRDSRGREWDKIEIDK"
RBS 4144..4152
/label=Shine-Dalgarno sequence
/note="full consensus sequence for ribosome-binding sites
upstream of start codons in E. coli; complementary to a
region in the 3' end of the 16S rRNA (Chen et al., 1994)"
CDS 4241..4954
/codon_start=1
/gene="cIts"
/product="temperature-sensitive variant of the phage lambda
repressor"
/label=lambda repressor (ts)
/note="thermosensitivity is conferred by the A67T mutation"
/translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
SGVGALFNGINALNAYNAALLTKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
PVFSHVQAGMFSPKLRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
VGKVIASQWPEETFG"
protein_bind complement(4961..4994)
/label=FRT (minimal)
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="supports FLP-mediated excision but not integration
(Turan and Bode, 2011)"
terminator 5071..5102
/label=tonB terminator
/note="bidirectional E. coli tonB-P14 transcription
terminator"
terminator complement(5151..5178)
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
misc_feature 5233..5253
/label=BioBrick suffix
/note="universal suffix for all parts"
rep_origin complement(5313..5901)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5393..5412)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5931..6236)
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind complement(6033..6052)
/label=ccdB-fwd
/note="ccdB gene, forward primer"
terminator complement(6468..6554)
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator complement(6651..6737)
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
ORIGIN
1 aattcctggc agtttatggc gggcgtcctg cccgccaccc tccgggccgt tgcttcgcaa
61 cgttcaaatc cgctcccggc ggatttgtcc tactcaggag agcgttcacc gacaaacaac
121 agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga tgcctgggga
181 attgggaatt aattcttgaa gacgaaaggg cctcgtgata cgcctatttt tataggttaa
241 tgtcatgtgc agcatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
301 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
361 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
421 aacgcgcgag gcagccggat caattctcat gtttgacagc ttatcactga tcagtgaatt
481 aatggcgatg acgcatcctc acgataatat ccgggtaggc gcaatcactt tcgtctctac
541 tccgttacaa agcgaggctg ggtatttccc ggcctttctg ttatccgaaa tccactgaaa
601 gcacagcggc tggctgagga gataaataat aaacgagggg ctgtatgcac aaagcatctt
661 ctgttgagtt aagaacgagt atcgagatgg cacatagcct tgctcaaatt ggaatcaggt
721 ttgtgccaat accagtagaa acagacgaag aagctagcta atgctctgtc tcaggtcact
781 aatactatct aagtagttga ttcatagtga ctggatatgt tgcgttttgt cgcattatgt
841 agtctatcat ttaaccacag attagtgtaa tgcgatgatt tttaagtgat taatgttatt
901 ttgtcatcct ttaggtgaat aagttgtata tttaaaatct ctttaattat cagtaaatta
961 atgtaagtag gtcattatta gtcaaaataa aatcatttgt cgatttcaat tttgtcccat
1021 ggcatatgga agttcctata ctttctagag aataggaact tcggaatagg aacttcctaa
1081 ttcccatgtc agccgttaag tgttcctgtg tcactcaaaa ttgctttgag aggctctaag
1141 ggcttctcag tgcgttacat ccctggcttg ttgtccacaa ccgttaaacc ttaaaagctt
1201 taaaagcctt atatattctt ttttttctta taaaacttaa aaccttagag gctatttaag
1261 ttgctgattt atattaattt tattgttcaa acatgagagc ttagtacgtg aaacatgaga
1321 gcttagtacg ttagccatga gagcttagta cgttagccat gagggtttag ttcgttaaac
1381 atgagagctt agtacgttaa acatgagagc ttagtacgtg aaacatgaga gcttagtacg
1441 tactatcaac aggttgaact gctgatcttc agatcctcta cgccggacgc atcgtggccg
1501 gatcttgcgg ccgcactagg tcgacgcttg gattctcacc aataaaaaac gcccggcggc
1561 aaccgagcgt tctgaacaaa tccagatgga gttctgaggt cattactgga tctatcaaca
1621 ggagtccaag cgagctcaaa aattaaaaat gaagttttgg aggcctcatt tggtgacgaa
1681 ataactaagc acttgtctcc tgtttactcc cctgagcttg aggggtcaac atgaaggtca
1741 ttgatagcag gataataata cagtaaaacg ctaaaccaat aatccaaatc cagccatccc
1801 aaattggtag tgaatgatta taaataacag caaacagtaa tgggccaata acaccggttg
1861 cattggtaag gctcaccaat aatccctgta aagcaccttg ctgatgactc tttgtttgga
1921 tagacatcac tccctgtaat gcaggtaaag cgatcccacc accagccaat aaaattaaaa
1981 cagggaaaac taaccaacct tcagatataa acgctaaaaa ggcaaatgca ctactatctg
2041 caataaatcc gagcagtact gccgtttttt cgccccattt agtggctatt cttcctgcca
2101 caaaggcttg gaatactgag tgtaaaagac caagacccgc taatgaaaag ccaaccatca
2161 tgctattcca tccaaaacga ttttcggtaa atagcaccca caccgttgcg ggaatttggc
2221 ctatcaattg cgctgaaaaa taaataatca acaaaatggg catcgtttta aataaagtga
2281 tgtacaccga atttgattgc gtctcaaccc ctacttcggt atctgtatta tcacgtgtat
2341 ttttggtttc acggaaccaa aacataacca caaggaaagt gacaatattt agcaacgcag
2401 cgataaaaaa gggactatgc ggtgaaatct ctcctgcaaa accaccaata ataggccccg
2461 ctattaaacc aagcccaaaa cttgccccta accaaccgaa ccacttcacg cgttgagaag
2521 ctgaggtggt atcggcaatg accgatgccg cgacagcccc agtagctcct gtgatccctg
2581 aaagcaaacg gcctaaatac agcatccaaa gcgcacttga aaaagccagc aataagtaat
2641 ccagcgatgc gcctattaat gacaacaaca gcactgggcg ccgaccaaat cggtcagaca
2701 tttttccaag ccaaggagca aagataacct gcattaacgc ataaagtgca agcaatacgc
2761 caaagtggtt agcgatatct tccgaagcaa taaattcacg taataacgtt ggcaagactg
2821 gcatgataag gccaatcccc atggcatcga gcaacgtaat taccaatgcg atctttgtcg
2881 aactattcat ttcacttttc tctatcactg atagggagtg ggaaaataac tctatcaatg
2941 atagggtgtc aaatcgatgc acaacgtgcg ttacatatca aagggaaaac tgtccatacc
3001 catggtaagt aggtcattat tagtcaaaat aaaatcattt gtcgatttca attttgtccc
3061 attcccgtcc acggctgtcc ctataccgcg ccgccattga atctgattta tgcccgagaa
3121 gacgttgagc aaacttatcg ccaatctggt tccggtatag cctcgctgac aggctacgca
3181 gttcatggaa tgttggcggg tttccatcaa atgagagtcc agatgcattt ctcgcctttg
3241 taaaatactt tgatactgtt ttcggggaaa gcggatcgtg atgctttgat gcgattatag
3301 tttcactgct gctggcctcc ctgcattgct gtagtgtatc agccaatgag atattgagcg
3361 cgtcaatcgt tagcgttagt ggaatggcga gtttagcccc tgttttactc tgttcaatgt
3421 gaagatggtt gtcgtttatg tctgaccatt tcattctgca caaatcgcct actctctgcc
3481 ctgtaacgac agccaaatcc attgccagtc tcagccagat tgggagcggt tcggctgcat
3541 ggtatatcgc gacatattca ttagctgtca gtcttgaacg ccttacttct gattttgctg
3601 tgcgggttgc tgttaccgga ttcgtagcca catgcccctc ggctattgcc tcacgaaaaa
3661 cgtcaacaag ggttgacctg atcaattttg cggaagccga tttgccttct gctacgtagg
3721 tgtttagcat tgctgccacc tctttcgttg atatgtcagc gagcggtttg tccggcaatt
3781 ttcttcggat tgctctgatt ttgctggcgt agtcgagtag agttttcggc ctgatacccc
3841 tctcgctgag gattgtttca tatcggtcaa gccacgcatg aagagtgatt gcgtcagcgc
3901 ctttaattct gtctatcagt gactcacgcc tgttcccgga tagcaactca atgttggcct
3961 gtatagcttc agtgattgcg attcgcctgt ctcggcctaa tccaaactct ttacccgtcc
4021 ttgggtccct gtagcagtaa tatccattgt ttcttatata aaggttaggg ggtaaatccc
4081 ggcgctcatg acttcgcctt cttcccattt ctgatcctct tcaaaaggct acacgcgtat
4141 acaacctcct tagtacatgc aaccattatc accgccagag gtaaaatagt caacacgcac
4201 ggtgttagat atttatccct tgcggtgata gatttaacgt atgagcacaa aaaagaaacc
4261 attaacacaa gagcagcttg aggacgcacg tcgccttaaa gcaatttatg aaaaaaagaa
4321 aaatgaactt ggcttatccc aggaatctgt cgcagacaag atggggatgg ggcagtcagg
4381 cgttggtgct ttatttaatg gcatcaatgc attaaatgct tataacgccg cattgcttac
4441 aaaaattctc aaagttagcg ttgaagaatt tagcccttca atcgccagag aaatctacga
4501 gatgtatgaa gcggttagta tgcagccgtc acttagaagt gagtatgagt accctgtttt
4561 ttctcatgtt caggcaggga tgttctcacc taagcttaga acctttacca aaggtgatgc
4621 ggagagatgg gtaagcacaa ccaaaaaagc cagtgattct gcattctggc ttgaggttga
4681 aggtaattcc atgaccgcac caacaggctc caagccaagc tttcctgacg gaatgttaat
4741 tctcgttgac cctgagcagg ctgttgagcc aggtgatttc tgcatagcca gacttggggg
4801 tgatgagttt accttcaaga aactgatcag ggatagcggt caggtgtttt tacaaccact
4861 aaacccacag tacccaatga tcccatgcaa tgagagttgt tccgttgtgg ggaaagttat
4921 cgctagtcag tggcctgaag agacgtttgg ctgacctagg gaagttccta tactttctag
4981 agaataggaa cttcggacgt ctaagaaacc attattatca tgagggccca ccggttaatt
5041 aacggcacca ccgaaattca gtaagcagaa agtcaaaagc ctccgaccgg aggcttttga
5101 ctattactca acaggtaagg cgcgaggttt tttcaattgt tcgaaacgca aaaaggccat
5161 ccgtcaggat ggccttctgc ttaatttgat gcctgtcagt ttaggttagg cgccatgcat
5221 ctcgaggcat gcctgcagcg gccgctacta gtaatgtgag caaaaggcca gcaaaaggcc
5281 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
5341 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
5401 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
5461 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
5521 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
5581 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
5641 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
5701 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
5761 tttggtatct gcgctctgct aaagccagtt accttcggaa aaagagttgg tagctcttga
5821 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagta gcagattacg
5881 cgcagaaaaa aaggatctca agaagatctt ttgatctttt ctacgggtta ttatattccc
5941 cagaacatca ggttaatggc gtttttgatg tcattttcgc ggtggctgag atcagccact
6001 tcttccccga taacggagac cggcacactg gccatatcgg tggtcatcat gcgccagctt
6061 tcatccccga tatgcaccac cgggtaaagt tcacgggaga ctttatctga cagcagacgt
6121 gcactggcca gggggatcac catccgtcgc ccgggcgtgt caataatatc actctgtaca
6181 tccacaaaca gacgataacg gctctctctt ttataggtgt aaaccttaaa ctgcatagca
6241 tacctctttt tgacatactt cgggtataca tatcagtata tattcttata ccgcaaaaat
6301 cagcgcgcaa atacgcatac tgttatctgg cttttagtaa gccctctaga ggatccccgg
6361 gtaccgagct cgaattccga tccccaattc ctggcagttt atggcgggcg tcctgcccgc
6421 caccctccgg gccgttgctt cgcaacgttc aaatccgctc ccggcggatt tgtcctactc
6481 aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc
6541 tttcgtttta tttgatgcct ggggaattaa ttcctggcag tttatggcgg gcgtcctgcc
6601 cgccaccctc cgggccgttg cttcgcaacg ttcaaatccg ctcccggcgg atttgtccta
6661 ctcaggagag cgttcaccga caaacaacag ataaaacgaa aggcccagtc tttcgactga
6721 gcctttcgtt ttatttgatg cctggggaat t
//
