LOCUS pOSIP-CH (CamR, HK022) 6224 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 6224)
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 6224)
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-CH (CamR, HK022)
FEATURES Location/Qualifiers
source 1..6224
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 142..447
/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 326..345
/label=ccdB-fwd
/note="ccdB gene, forward primer"
rep_origin 477..1065
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 966..985
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
misc_feature 1125..1145
/label=BioBrick suffix
/note="universal suffix for all parts"
terminator 1200..1227
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
terminator complement(1276..1307)
/label=tonB terminator
/note="bidirectional E. coli tonB-P14 transcription
terminator"
protein_bind 1384..1417
/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)"
CDS complement(1424..2137)
/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"
RBS 2226..2234
/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 2270..3343
/codon_start=1
/gene="int"
/product="integrase from phage HK022"
/label=HK022 integrase
/translation="MGRRRSHERRDLPPNLYIRNNGYYCYRDPRTGKEFGLGRDRRIAI
TEAIQANIELLSGNRRESLIDRIKGADAITLHAWLDRYETILSERGIRPKTLLDYASKI
RAIRRKLPDKPLADISTKEVAAMLNTYVAEGKSASAKLIRSTLVDVFREAIAEGHVATN
PVTATRTAKSEVRRSRLTANEYVAIYHAAEPLPIWLRLAMDLAVVTGQRVGDLCRMKWS
DINDNHLHIEQSKTGAKLAIPLTLTIDALNISLADTLQQCREASSSETIIASKHHDPLS
PKTVSKYFTKARNASGLSFDGNPPTFHELRSLSARLYRNQIGDKFAQRLLGHKSDSMAA
RYRDSRGREWDKIEIDK"
promoter 3442..3544
/label=cat promoter
/note="promoter of the E. coli cat gene encoding
chloramphenicol acetyltransferase"
CDS 3545..4204
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
primer_bind complement(3611..3630)
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
terminator 4225..4319
/label=lambda t0 terminator
/note="transcription terminator from phage lambda"
rep_origin 4376..4764
/label=R6K gamma ori
/note="gamma replication origin from E. coli plasmid R6K;
requires the R6K initiator protein pi for replication"
protein_bind 4775..4822
/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)."
protein_bind 4852..5078
/gene="attP"
/label=HK022 attP
/bound_moiety="HK022 integrase"
/note="attachment site of phage HK022"
terminator complement(5113..5359)
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
primer_bind complement(5450..5472)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 5623..5641
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
terminator 5682..5768
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 5865..5951
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 6048..6134
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
ORIGIN
1 gaattcgagc tcggtacccg gggatcctct agagggctta ctaaaagcca gataacagta
61 tgcgtatttg cgcgctgatt tttgcggtat aagaatatat actgatatgt atacccgaag
121 tatgtcaaaa agaggtatgc tatgcagttt aaggtttaca cctataaaag agagagccgt
181 tatcgtctgt ttgtggatgt acagagtgat attattgaca cgcccgggcg acggatggtg
241 atccccctgg ccagtgcacg tctgctgtca gataaagtct cccgtgaact ttacccggtg
301 gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag tgtgccggtc
361 tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat caaaaacgcc
421 attaacctga tgttctgggg aatataataa cccgtagaaa agatcaaaag atcttcttga
481 gatccttttt ttctgcgcgt aatctgctac ttgcaaacaa aaaaaccacc gctaccagcg
541 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggctttagc
601 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
661 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
721 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
781 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
841 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
901 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
961 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
1021 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
1081 gcctttttac ggttcctggc cttttgctgg ccttttgctc acattactag tagcggccgc
1141 tgcaggcatg cctcgagatg catggcgcct aacctaaact gacaggcatc aaattaagca
1201 gaaggccatc ctgacggatg gcctttttgc gtttcgaaca attgaaaaaa cctcgcgcct
1261 tacctgttga gtaatagtca aaagcctccg gtcggaggct tttgactttc tgcttactga
1321 atttcggtgg tgccgttaat taaccggtgg gccctcatga taataatggt ttcttagacg
1381 tccgaagttc ctattctcta gaaagtatag gaacttccct aggtcagcca aacgtctctt
1441 caggccactg actagcgata actttcccca caacggaaca actctcattg catgggatca
1501 ttgggtactg tgggtttagt ggttgtaaaa acacctgacc gctatccctg atcagtttct
1561 tgaaggtaaa ctcatcaccc ccaagtctgg ctatgcagaa atcacctggc tcaacagcct
1621 gctcagggtc aacgagaatt aacattccgt caggaaagct tggcttggag cctgttggtg
1681 cggtcatgga attaccttca acctcaagcc agaatgcaga atcactggct tttttggttg
1741 tgcttaccca tctctccgca tcacctttgg taaaggttct aagcttaggt gagaacatcc
1801 ctgcctgaac atgagaaaaa acagggtact catactcact tctaagtgac ggctgcatac
1861 taaccgcttc atacatctcg tagatttctc tggcgattga agggctaaat tcttcaacgc
1921 taactttgag aatttttgta agcaatgcgg cgttataagc atttaatgca ttgatgccat
1981 taaataaagc accaacgcct gactgcccca tccccatctt gtctgcgaca gattcctggg
2041 ataagccaag ttcatttttc tttttttcat aaattgcttt aaggcgacgt gcgtcctcaa
2101 gctgctcttg tgttaatggt ttcttttttg tgctcatacg ttaaatctat caccgcaagg
2161 gataaatatc taacaccgtg cgtgttgact attttacctc tggcggtgat aatggttgca
2221 tgtactaagg aggttgtata cgcgtgtagc cttttgaaga ggatcagaaa tgggaagaag
2281 gcgaagtcat gagcgccggg atttaccccc taacctttat ataagaaaca atggatatta
2341 ctgctacagg gacccaagga cgggtaaaga gtttggatta ggccgagaca ggcgaatcgc
2401 aatcactgaa gctatacagg ccaacattga gttgctatcc gggaacaggc gtgagtcact
2461 gatagacaga attaaaggcg ctgacgcaat cactcttcat gcgtggcttg accgatatga
2521 aacaatcctc agcgagaggg gtatcaggcc gaaaactcta ctcgactacg ccagcaaaat
2581 cagagcaatc cgaagaaaat tgccggacaa accgctcgct gacatatcaa cgaaagaggt
2641 ggcagcaatg ctaaacacct acgtagcaga aggcaaatcg gcttccgcaa aattgatcag
2701 gtcaaccctt gttgacgttt ttcgtgaggc aatagccgag gggcatgtgg ctacgaatcc
2761 ggtaacagca acccgcacag caaaatcaga agtaaggcgt tcaagactga cagctaatga
2821 atatgtcgcg atataccatg cagccgaacc gctcccaatc tggctgagac tggcaatgga
2881 tttggctgtc gttacagggc agagagtagg cgatttgtgc agaatgaaat ggtcagacat
2941 aaacgacaac catcttcaca ttgaacagag taaaacaggg gctaaactcg ccattccact
3001 aacgctaacg attgacgcgc tcaatatctc attggctgat acactacagc aatgcaggga
3061 ggccagcagc agtgaaacta taatcgcatc aaagcatcac gatccgcttt ccccgaaaac
3121 agtatcaaag tattttacaa aggcgagaaa tgcatctgga ctctcatttg atggaaaccc
3181 gccaacattc catgaactgc gtagcctgtc agcgaggcta taccggaacc agattggcga
3241 taagtttgct caacgtcttc tcgggcataa atcagattca atggcggcgc ggtataggga
3301 cagccgtgga cgggaatggg acaaaattga aatcgacaaa tgattttatt ttgactaata
3361 atgacctact taccatgggt atggacagtt ttccctttga tatgtaacgc acgttgtgga
3421 tatctggcga aaatgagacg ttgatcggca cgtaagaggt tccaactttc accataatga
3481 aataagatca ctaccgggcg tattttttga gttatcgaga ttttcaggag ctaaggaagc
3541 taaaatggag aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa
3601 agaacatttt gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct
3661 ggatattacg gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt
3721 tattcacatt cttgcccgcc tgatgaatgc tcatccggaa ttccgtatgg caatgaaaga
3781 cggtgagctg gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac
3841 tgaaacgttt tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat
3901 atattcgcaa gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat
3961 tgagaatatg tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa
4021 cgtggccaat atggacaact tcttcgcccc cgttttcacc atgggcaaat attatacgca
4081 aggcgacaag gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatggctt
4141 ccatgtcggc agaatgctta atgaattaca acagtactgc gatgagtggc agggcggggc
4201 gtaatttgat atcgagctcg cttggactcc tgttgataga tccagtaatg acctcagaac
4261 tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt ggtgagaatc
4321 caagcgtcga cctagtgcgg ccgcaagatc cggccacgat gcgtccggcg tagaggatct
4381 gaagatcagc agttcaacct gttgatagta cgtactaagc tctcatgttt cacgtactaa
4441 gctctcatgt ttaacgtact aagctctcat gtttaacgaa ctaaaccctc atggctaacg
4501 tactaagctc tcatggctaa cgtactaagc tctcatgttt cacgtactaa gctctcatgt
4561 ttgaacaata aaattaatat aaatcagcaa cttaaatagc ctctaaggtt ttaagtttta
4621 taagaaaaaa aagaatatat aaggctttta aagcttttaa ggtttaacgg ttgtggacaa
4681 caagccaggg atgtaacgca ctgagaagcc cttagagcct ctcaaagcaa ttttgagtga
4741 cacaggaaca cttaacggct gacatgggaa ttaggaagtt cctattccga agttcctatt
4801 ctctagaaag tataggaact tccatatgcc atgggacaaa attgaaatcg acaaatgatt
4861 ttattttgac taataatgac ctacttacat taatttactg ataattaaag agattttaaa
4921 tatacaactt attcacctaa aggatgacaa aataacatta atcacttaaa aatcatcgca
4981 ttacactaat ctgtggttaa atgatagact acataatgcg acaaaacgca acatatccag
5041 tcactatgaa tcaactactt agatagtatt agtgacctga gacagagcat tagctagctt
5101 cttcgtctgt ttctactggt attggcacaa acctgattcc aatttgagca aggctatgtg
5161 ccatctcgat actcgttctt aactcaacag aagatgcttt gtgcatacag cccctcgttt
5221 attatttatc tcctcagcca gccgctgtgc tttcagtgga tttcggataa cagaaaggcc
5281 gggaaatacc cagcctcgct ttgtaacgga gtagagacga aagtgattgc gcctacccgg
5341 atattatcgt gaggatgcgt catcgccatt aattcactga tcagtgataa gctgtcaaac
5401 atgagaattg atccggctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
5461 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
5521 gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta
5581 gcgatagcgg agtgtatgct gcacatgaca ttaacctata aaaataggcg tatcacgagg
5641 ccctttcgtc ttcaagaatt aattcccaat tccccaggca tcaaataaaa cgaaaggctc
5701 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
5761 ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg
5821 caggacgccc gccataaact gccaggaatt aattccccag gcatcaaata aaacgaaagg
5881 ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga
5941 gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc
6001 gggcaggacg cccgccataa actgccagga attaattccc caggcatcaa ataaaacgaa
6061 aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc
6121 tgagtaggac aaatccgccg ggagcggatt tgaacgttgc gaagcaacgg cccggagggt
6181 ggcgggcagg acgcccgcca taaactgcca ggaattgggg atcg
//
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