LOCUS pOSIP-KL (KanR, Lambda) 6406 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
6406)
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 6406)
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-KL (KanR, Lambda)
FEATURES Location/Qualifiers
source
1..6406
/mol_type="other DNA"
/organism="synthetic DNA construct"
terminator 115..201
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
CDS 433..738
/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
617..636
/label=ccdB-fwd
/note="ccdB gene,
forward primer"
rep_origin 768..1356
/direction=RIGHT
/label=ori
/note="high-copy-number
ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind
1257..1276
/label=pBR322ori-F
/note="pBR322
origin, forward primer"
misc_feature 1416..1436
/label=BioBrick suffix
/note="universal suffix for all parts"
terminator 1491..1518
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
terminator complement(1567..1598)
/label=tonB
terminator
/note="bidirectional E. coli tonB-P14 transcription
terminator"
protein_bind 1675..1708
/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(1714..2427)
/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 2516..2524
/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 2669..3739
/codon_start=1
/gene="int"
/product="integrase from phage
lambda"
/label=lambda integrase
/translation="MGRRRSHERRDLPPNLYIRNNGYYCYRDPRTGKEFGLGRDRRIAI
TEAIQANIELFSGHKHKPLTARINSDNSVTLHSWLDRYEKILASRGIKQKTLINYMSKI
KAIRRGLPDAPLEDITTKEIAAMLNGYIDEGKAASAKLIRSTLSDAFREAIAEGHITTN
HVAATRAAKSEVRRSRLTADEYLKIYQAAESSPCWLRLAMELAVVTGQRVGDLCEMKWS
DIVDGYLYVEQSKTGVKIAIPTALHIDALGISMKETLDKCKEILGGETIIASTRREPLS
SGTVSRYFMRARKASGLSFEGDPPTFHELRSLSARLYEKQISDKFAQHLLGHKSDTMAS
QYRDDRGREWDKIEIK"
primer_bind complement(3955..3974)
/label=pENTR-R
/note="pENTR vectors, reverse primer"
CDS
3966..4781
/codon_start=1
/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 complement
(4039..4058)
/label=Kan-R
/note="Kanamycin
resistance gene, reverse primer"
rep_origin 4903..5291
/label=R6K gamma ori
/note="gamma replication origin from E.
coli plasmid R6K;
requires the R6K initiator protein pi for
replication"
protein_bind 5302..5349
/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 5412..5645
/gene="attP"
/label=lambda attP
/bound_moiety="lambda integrase"
/note="integrase from phage lambda"
terminator complement
(5769..6015)
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
primer_bind
complement(6106..6128)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 6279..6297
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI
site, forward
primer"
terminator join
(6338..6406,1..18)
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1
from the E. coli rrnB
gene"
ORIGIN
1 ctcctgagta
ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga
61 gggtggcggg
caggacgccc gccataaact gccaggaatt aattccccag gcatcaaata
121 aaacgaaagg
ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
181 gctctcctga
gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
241 ggagggtggc
gggcaggacg cccgccataa actgccagga attggggatc ggaattcgag
301 ctcggtaccc
ggggatcctc tagagggctt actaaaagcc agataacagt atgcgtattt
361 gcgcgctgat
ttttgcggta taagaatata tactgatatg tatacccgaa gtatgtcaaa
421 aagaggtatg
ctatgcagtt taaggtttac acctataaaa gagagagccg ttatcgtctg
481 tttgtggatg
tacagagtga tattattgac acgcccgggc gacggatggt gatccccctg
541 gccagtgcac
gtctgctgtc agataaagtc tcccgtgaac tttacccggt ggtgcatatc
601 ggggatgaaa
gctggcgcat gatgaccacc gatatggcca gtgtgccggt ctccgttatc
661 ggggaagaag
tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg
721 atgttctggg
gaatataata acccgtagaa aagatcaaaa gatcttcttg agatcctttt
781 tttctgcgcg
taatctgcta cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
841 ttgccggatc
aagagctacc aactcttttt ccgaaggtaa ctggctttag cagagcgcag
901 ataccaaata
ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta
961 gcaccgccta
catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
1021 aagtcgtgtc
ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
1081 ggctgaacgg
ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
1141 agatacctac
agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
1201 aggtatccgg
taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
1261 aacgcctggt
atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
1321 ttgtgatgct
cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
1381 cggttcctgg
ccttttgctg gccttttgct cacattacta gtagcggccg ctgcaggcat
1441 gcctcgagat
gcatggcgcc taacctaaac tgacaggcat caaattaagc agaaggccat
1501 cctgacggat
ggcctttttg cgtttcgaac aattgaaaaa acctcgcgcc ttacctgttg
1561 agtaatagtc
aaaagcctcc ggtcggaggc ttttgacttt ctgcttactg aatttcggtg
1621 gtgccgttaa
ttaaccggtg ggccctcatg ataataatgg tttcttagac gtccgaagtt
1681 cctattctct
agaaagtata ggaacttccc tagtcagcca aacgtctctt caggccactg
1741 actagcgata
actttcccca caacggaaca actctcattg catgggatca ttgggtactg
1801 tgggtttagt
ggttgtaaaa acacctgacc gctatccctg atcagtttct tgaaggtaaa
1861 ctcatcaccc
ccaagtctgg ctatgcagaa atcacctggc tcaacagcct gctcagggtc
1921 aacgagaatt
aacattccgt caggaaagct tggcttggag cctgttggtg cggtcatgga
1981 attaccttca
acctcaagcc agaatgcaga atcactggct tttttggttg tgcttaccca
2041 tctctccgca
tcacctttgg taaaggttct aagcttaggt gagaacatcc ctgcctgaac
2101 atgagaaaaa
acagggtact catactcact tctaagtgac ggctgcatac taaccgcttc
2161 atacatctcg
tagatttctc tggcgattga agggctaaat tcttcaacgc taactttgag
2221 aatttttgta
agcaatgcgg cgttataagc atttaatgca ttgatgccat taaataaagc
2281 accaacgcct
gactgcccca tccccatctt gtctgcgaca gattcctggg ataagccaag
2341 ttcatttttc
tttttttcat aaattgcttt aaggcgacgt gcgtcctcaa gctgctcttg
2401 tgttaatggt
ttcttttttg tgctcatacg ttaaatctat caccgcaagg gataaatatc
2461 taacaccgtg
cgtgttgact attttacctc tggcggtgat aatggttgca tgtactaagg
2521 aggttgtata
cgcgtcgtcg atgggttcgg gaatgcagga tattcccacc tccggttaag
2581 gatggaagag
agtatctgtt ccacgaatca gcggtaaagg ttgacttaaa tcgaccagta
2641 acaggtggcc
ttttgaagag gatcagaaat gggaagaagg cgaagtcatg agcgccggga
2701 tttaccccct
aacctttata taagaaacaa tggatattac tgctacaggg acccaaggac
2761 gggtaaagag
tttggattag gcagagacag gcgaatcgca atcactgaag ctatacaggc
2821 caacattgag
ttattttcag gacacaaaca caagcctctg acagcgagaa tcaacagtga
2881 taattccgtt
acgttacatt catggcttga tcgctacgaa aaaatcctgg ccagcagagg
2941 aatcaagcag
aagacactca taaattacat gagcaaaatt aaagcaataa ggaggggtct
3001 gcctgatgct
ccacttgaag acatcaccac aaaagaaatt gcggcaatgc tcaatggata
3061 catagacgag
ggcaaggcgg cgtcagccaa gttaatcaga tcaacactga gcgatgcatt
3121 ccgagaggca
atagctgaag gccatataac aacaaaccat gtcgctgcca ctcgcgcagc
3181 aaaatcagag
gtaaggagat caagacttac ggctgacgaa tacctgaaaa tttatcaagc
3241 agcagaatca
tcaccatgtt ggctcagact tgcaatggaa ctggctgttg ttaccgggca
3301 acgagttggt
gatttatgcg aaatgaagtg gtctgatatc gtagatggat atctttatgt
3361 cgagcaaagc
aaaacaggcg taaaaattgc catcccaaca gcattgcata ttgatgctct
3421 cggaatatca
atgaaggaaa cacttgataa atgcaaagag attcttggcg gagaaaccat
3481 aattgcatct
actcgtcgcg aaccgctttc atccggcaca gtatcaaggt attttatgcg
3541 cgcacgaaaa
gcatcaggtc tttccttcga aggggatccg cctacctttc acgagttgcg
3601 cagtttgtct
gcaagactct atgagaagca gataagcgat aagtttgctc aacatcttct
3661 cgggcataag
tcggacacca tggcatcaca gtatcgtgat gacagaggca gggagtggga
3721 caaaattgaa
atcaaataat gattttattt tgactgatag tgacctgttc gttgcaacaa
3781 attgataagc
aatgccaagc ttggcactgg ctgatcagct agcccatggg tatggacagt
3841 tttccctttg
atatgtaacg cacgttgtgt ctcaaaatct ctgatgttac attgcacaag
3901 ataaaaatat
atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
3961 gtgttatgag
ccatattcaa cgggaaacgt cttgctcccg tccgcgctta aactccaaca
4021 tggacgctga
tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
4081 caatctatcg
cttgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
4141 gtagcgttgc
caatgatgtt acagatgaga tggtccgtct caactggctg acggagttta
4201 tgcctctccc
gaccatcaag cattttatcc gtactcctga tgatgcgtgg ttactcacca
4261 ccgcgattcc
tgggaaaaca gccttccagg tattagaaga atatcctgat tcaggtgaaa
4321 atattgttga
tgcgctggcc gtgttcctgc gccggttaca ttcgattcct gtttgtaatt
4381 gtccttttaa
cagcgatcgt gtatttcgtc ttgctcaggc gcaatcacgc atgaataacg
4441 gtttggttga
tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
4501 ggaaagaaat
gcacaagctc ttgccattct caccggattc agtcgtcact catggtgatt
4561 tctcacttga
taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
4621 gggtcggaat
cgcagaccgt taccaggacc ttgccattct ttggaactgc ctcggtgagt
4681 tttctccttc
attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
4741 ataaattgca
gtttcatttg atgctcgatg agtttttcta ataatactag cagaaatcat
4801 ccttagcgaa
agctaaggat tttttttatc tgattaccgc ctttgagtga gcgtcgacct
4861 agtgcggccg
caagatccgg ccacgatgcg tccggcgtag aggatctgaa gatcagcagt
4921 tcaacctgtt
gatagtacgt actaagctct catgtttcac gtactaagct ctcatgttta
4981 acgtactaag
ctctcatgtt taacgaacta aaccctcatg gctaacgtac taagctctca
5041 tggctaacgt
actaagctct catgtttcac gtactaagct ctcatgtttg aacaataaaa
5101 ttaatataaa
tcagcaactt aaatagcctc taaggtttta agttttataa gaaaaaaaag
5161 aatatataag
gcttttaaag cttttaaggt ttaacggttg tggacaacaa gccagggatg
5221 taacgcactg
agaagccctt agagcctctc aaagcaattt tgagtgacac aggaacactt
5281 aacggctgac
atgggaatta ggaagttcct attccgaagt tcctattctc tagaaagtat
5341 aggaacttcc
atatgccatg gcatcacagt atcgtgatga cagaggcagg gagtgggaca
5401 aaattgaaat
caaataatga ttttattttg actgatagtg acctgttcgt tgcaacaaat
5461 tgataagcaa
tgctttttta taatgccaac ttagtataaa aaagctgaac gagaaacgta
5521 aaatgatata
aatatcaata tattaaatta gattttgcat aaaaaacaga ctacataata
5581 ctgtaaaaca
caacatatat gcagtcacta tgaatcaact acttagatgg tattagtgac
5641 ctgtaacaga
gcattagcgc aaggtgattt ttgtcttctt gcgctaattt tttgtcatca
5701 aacctgtcgc
actccagaga agcacaaagc ctcgcaatcc agtgcaaagc tagcttcttc
5761 gtctgtttct
actggtattg gcacaaacct gattccaatt tgagcaaggc tatgtgccat
5821 ctcgatactc
gttcttaact caacagaaga tgctttgtgc atacagcccc tcgtttatta
5881 tttatctcct
cagccagccg ctgtgctttc agtggatttc ggataacaga aaggccggga
5941 aatacccagc
ctcgctttgt aacggagtag agacgaaagt gattgcgcct acccggatat
6001 tatcgtgagg
atgcgtcatc gccattaatt cactgatcag tgataagctg tcaaacatga
6061 gaattgatcc
ggctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
6121 gctcccggag
acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
6181 gggcgcgtca
gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
6241 tagcggagtg
tatgctgcac atgacattaa cctataaaaa taggcgtatc acgaggccct
6301 ttcgtcttca
agaattaatt cccaattccc caggcatcaa ataaaacgaa aggctcagtc
6361 gaaagactgg
gcctttcgtt ttatctgttg tttgtcggtg aacgct
//
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