LOCUS pOSIP-CT (CamR, P21) 6694 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 6694)
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 6694)
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-CT (CamR, P21)
FEATURES Location/Qualifiers
source 1..6694
/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 2299..3441
/codon_start=1
/gene="int"
/product="integrase from phage phi-21"
/label=phage phi-21 integrase
/translation="MAARPRSHKISIPNLYCKLDKRTGKVYWQYKHPLSGRFHSLGTDE
NEAKQVATEANTIIAEQRTRQILSVNERLERMKGRRSDITVTEWLDKYISIQEDRLQHN
ELRPNSYRQKGKPIRLFREHCGMQHLKDITALDIAEIIDAVKAEGHNRMAQVVRMVLID
VFKEAQHAGHVPPGFNPAQATKQPRNRVNRQRLSLLEWQAIFESVSRRQPYLKCGMLLA
LVTGQRLGDICNLKFSDIWDDMLHITQEKTGSKLAIPLNLKCDALNITLREVISQCRDA
VVSKYLVHYRHTTSQANRGDQVSANTLTTAFKKAKEKCGIKWEQGTAPTFHEQRSLSER
LYREQGLDTQKLLGHKSRKMTDRYNDDRGKDWIIVDIKTA"
promoter 3603..3705
/label=cat promoter
/note="promoter of the E. coli cat gene encoding
chloramphenicol acetyltransferase"
CDS 3706..4365
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
primer_bind complement(3772..3791)
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
terminator 4386..4480
/label=lambda t0 terminator
/note="transcription terminator from phage lambda"
rep_origin 4537..4925
/label=R6K gamma ori
/note="gamma replication origin from E. coli plasmid R6K;
requires the R6K initiator protein pi for replication"
protein_bind 4936..4983
/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 5063..5562
/label=phage phi-21 attP
/bound_moiety="phage phi-21 integrase"
/note="attachment site of phage phi-21"
terminator complement(5583..5829)
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
primer_bind complement(5920..5942)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 6093..6111
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
terminator 6152..6238
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 6335..6421
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 6518..6604
/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 cgcgtagaac cgcaactccc aataaacgca aacccaaaac
2281 tccaacggat aatcgctgat ggctgctaga ccccgatctc acaaaatctc tatacccaat
2341 ttatattgca aattagataa gcgaaccgga aaggtatatt ggcaatacaa acatccacta
2401 tccggtcgtt ttcatagctt aggaactgat gagaatgaag caaaacaagt tgctactgaa
2461 gcaaatacca ttattgctga acaacgtacc agacaaatat taagcgtcaa tgagcgtctg
2521 gaaagaatga aaggcaggcg ctcagacatt acggtgacag aatggcttga taaatatatt
2581 tctatccagg aggacaggct gcaacataat gaactaagac ccaactccta tcggcaaaaa
2641 ggcaaaccca ttcgtctttt ccgtgagcat tgtggaatgc aacacctcaa ggatattacc
2701 gcacttgata ttgccgaaat aattgatgct gtaaaggctg aaggtcataa caggatggcg
2761 caagtcgtga gaatggtgtt gatcgacgtc ttcaaagaag cacaacacgc aggacatgtt
2821 ccgccaggat ttaacccagc gcaggcaaca aaacaaccgc gaaatcgagt aaaccgccaa
2881 agattgtcac tgctcgaatg gcaggcaata tttgaaagcg taagcagacg gcagccctat
2941 ttaaaatgcg gcatgctact tgctcttgtt actggacaac gtttaggcga tatctgcaat
3001 ttgaaattct ctgatatatg ggacgacatg ttgcacatta ctcaggaaaa aaccggttca
3061 aaacttgcta ttccgcttaa cctgaaatgc gatgctctga atattaccct tcgtgaagtt
3121 atatctcagt gcagggatgc tgttgttagt aaatatctgg tccattaccg tcacactacc
3181 tctcaagcaa acagaggaga ccaggtttct gcaaatactc tgacaacggc ttttaaaaag
3241 gccaaggaaa aatgtggcat aaaatgggag caaggaactg cgcccacatt tcatgaacag
3301 cgatctctgt cagaacgatt atatcgggaa cagggtctgg atacgcaaaa gttgttaggc
3361 cataaatcca gaaaaatgac cgaccgatac aatgatgatc gtggtaaaga ctggattatc
3421 gtagatatca aaacagcata gaaaatagcc agttttgggg aagggttttg gggaaagttt
3481 tggggaagat tttacatcat cataaaacaa cgggcgtgtt atacgcccgt tatccatggg
3541 tatggacagt tttccctttg atatgtaacg cacgttgtgg atatctggcg aaaatgagac
3601 gttgatcggc acgtaagagg ttccaacttt caccataatg aaataagatc actaccgggc
3661 gtattttttg agttatcgag attttcagga gctaaggaag ctaaaatgga gaaaaaaatc
3721 actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt tgaggcattt
3781 cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac ggccttttta
3841 aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat tcttgcccgc
3901 ctgatgaatg ctcatccgga attccgtatg gcaatgaaag acggtgagct ggtgatatgg
3961 gatagtgttc acccttgtta caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc
4021 tggagtgaat accacgacga tttccggcag tttctacaca tatattcgca agatgtggcg
4081 tgttacggtg aaaacctggc ctatttccct aaagggttta ttgagaatat gtttttcgtc
4141 tcagccaatc cctgggtgag tttcaccagt tttgatttaa acgtggccaa tatggacaac
4201 ttcttcgccc ccgttttcac catgggcaaa tattatacgc aaggcgacaa ggtgctgatg
4261 ccgctggcga ttcaggttca tcatgccgtt tgtgatggct tccatgtcgg cagaatgctt
4321 aatgaattac aacagtactg cgatgagtgg cagggcgggg cgtaatttga tatcgagctc
4381 gcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg atttgttcag
4441 aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaagcgtcg acctagtgcg
4501 gccgcaagat ccggccacga tgcgtccggc gtagaggatc tgaagatcag cagttcaacc
4561 tgttgatagt acgtactaag ctctcatgtt tcacgtacta agctctcatg tttaacgtac
4621 taagctctca tgtttaacga actaaaccct catggctaac gtactaagct ctcatggcta
4681 acgtactaag ctctcatgtt tcacgtacta agctctcatg tttgaacaat aaaattaata
4741 taaatcagca acttaaatag cctctaaggt tttaagtttt ataagaaaaa aaagaatata
4801 taaggctttt aaagctttta aggtttaacg gttgtggaca acaagccagg gatgtaacgc
4861 actgagaagc ccttagagcc tctcaaagca attttgagtg acacaggaac acttaacggc
4921 tgacatggga attaggaagt tcctattccg aagttcctat tctctagaaa gtataggaac
4981 ttccatatgc catggaatga ccgaccgata caatgatgat cgtggtaaag actggattat
5041 cgtagatatc aaaacagcat agaaaatagc cagttttggg gaagggtttt ggggaaagtt
5101 ttggggaaga ttttacatca tcataaaaca acgggcgtgt tatacgcccg ttataatatt
5161 taacacatgt ggtaattaca tatttttgat gatcgcgtca ccaaactcgc tgcatttcag
5221 cagtttagcg ccttccatca gacgttcgaa gtcataggtt acggtcttgg cgttgattgc
5281 gccttccata cctttaacaa tcaggtctgc ggcttcagtc cagcccatat ggcgcagcag
5341 tatataaaaa ttatgtattc aactcgctga tatttaatca taaatattat tttcttgtct
5401 ttgtattagc caaaatcaac tttattcaac cgtttgattt cattgacgtt cgtttttatt
5461 ttggggaaag attttctcta ccatttatat caaaattaca agtctacatt ttattaacat
5521 ctcaatacat gtgtggcctt gcaagccatt gcgaggcctt atgctagctt cttcgtctgt
5581 ttctactggt attggcacaa acctgattcc aatttgagca aggctatgtg ccatctcgat
5641 actcgttctt aactcaacag aagatgcttt gtgcatacag cccctcgttt attatttatc
5701 tcctcagcca gccgctgtgc tttcagtgga tttcggataa cagaaaggcc gggaaatacc
5761 cagcctcgct ttgtaacgga gtagagacga aagtgattgc gcctacccgg atattatcgt
5821 gaggatgcgt catcgccatt aattcactga tcagtgataa gctgtcaaac atgagaattg
5881 atccggctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
5941 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
6001 gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg
6061 agtgtatgct gcacatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
6121 ttcaagaatt aattcccaat tccccaggca tcaaataaaa cgaaaggctc agtcgaaaga
6181 ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc
6241 gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg caggacgccc
6301 gccataaact gccaggaatt aattccccag gcatcaaata aaacgaaagg ctcagtcgaa
6361 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa
6421 tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc gggcaggacg
6481 cccgccataa actgccagga attaattccc caggcatcaa ataaaacgaa aggctcagtc
6541 gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac
6601 aaatccgccg ggagcggatt tgaacgttgc gaagcaacgg cccggagggt ggcgggcagg
6661 acgcccgcca taaactgcca ggaattgggg atcg
//
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