LOCUS       sequence-53662-e        5626 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5626)
  AUTHORS   Kittleson JT, DeLoache W, Cheng HY, Anderson JC
  TITLE     Scalable plasmid transfer using engineered P1-based phagemids.
  JOURNAL   ACS Synth Biol. 2012 Dec 21;1(12):583-9. doi: 10.1021/sb300054p. 
            Epub 2012 Aug 30.
  PUBMED    23656280
REFERENCE   2  (bases 1 to 5626)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1021/sb300054p";
            journalName: "ACS Synth Biol"; date: "2012-12-21- 21"; volume: "1"; 
            issue: "12"; pages: "583-9"
COMMENT     Sequence Label: BBa_J72110-BBa_J72097
FEATURES             Location/Qualifiers
     source          1..5626
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(25..873)
                     /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(947..1783)
                     /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        1929..2047
                     /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)"
     terminator      2312..2383
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2399..2426
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    complement(2538..2562)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     rep_origin      2642..3187
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     3088..3107
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             3610..4269
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3676..3695)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      4290..4384
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     protein_bind    complement(4397..4430)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4468..4572
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4573..4641,4642..5433)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4791..4810)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     protein_bind    5533..5557
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      complement(5581..5624)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 gaattcatga gatctcatgg cgaattaatc tttctgcgaa ttgagatgac gccactggct
       61 gggcgtcatc ccggtttccc gggtaaacac caccgaaaaa tagttactat cttcaaagcc
      121 acattcggtc gaaatatcac tgattaacag gcggctatgc tggagaagat attgcgcatg
      181 acacactctg acctgtcgca gatattgatt gatggtcatt ccagtctgct ggcgaaattg
      241 ctgacgcaaa acgcgctcac tgcacgatgc ctcatcacaa aatttatcca gcgcaaaggg
      301 acttttcagg ctagccgcca gccgggtaat cagcttatcc agcaacgttt cgctggatgt
      361 tggcggcaac gaatcactgg tgtaacgatg gcgattcagc aacatcacca actgcccgaa
      421 cagcaactca gccatttcgt tagcaaacgg cacatgctga ctactttcat gctcaagctg
      481 accgataacc tgccgcgcct gcgccatccc catgctacct aagcgccagt gtggttgccc
      541 tgcgctggcg ttaaatcccg gaatcgcccc ctgccagtca agattcagtt tcagacgctc
      601 cgggcaataa ataatattct gcaaaaccag atcgttaacg gaagcgtagg agtgtttatc
      661 gtcagcatga atgtaaaaga gatcgccacg ggtaatgcga taagggcgat cgttgagtac
      721 atgcaggcca ttaccgcgcc agacaatcac cagctcacaa aaatcatgtg tatgttcagc
      781 aaagacatct tgcggataac ggtcagccac agcgactgcc tgctggtcgc tggcaaaaaa
      841 atcatctttg agaagtttta actgatgcgc caccgtggct acctcggcca gagaacgaag
      901 ttgattattc gcaatatggc gtacaaatac gttgagaaga ttcgcgttat tgcagaaagc
      961 catcccgtcc ctggcgaata tcacgcggtg accagttaaa ctctcggcga aaaagcgtcg
     1021 aaaagtggtt actgtcgctg aatccacagc gataggcgat gtcagtaacg ctggcctcgc
     1081 tgtggcgtag cagatgtcgg gctttcatca gtcgcaggcg gttcaggtat cgctgaggcg
     1141 tcagtcccgt ttgctgctta agctgccgat gtagcgtacg cagtgaaaga gaaaattgat
     1201 ccgccacggc atcccaattc acctcatcgg caaaatggtc ctccagccag gccagaagca
     1261 agttgagacg tgatgcgctg ttttccaggt tctcctgcaa actgctttta cgcagcaaga
     1321 gcagtaattg cataaacaag atttcgcgac tggcggtcga gggtaaatca ttttcccctt
     1381 cctgctgttc catctgtgca accagctgtc gcacctgctg caatacgctg tggttaacgc
     1441 gccagtgaga cggatactgc ccatccagct cttgtggcag caactgattc agcccggcga
     1501 gaaactgaaa tcgatccggc gagcgataca gcacattggt cagacacaga ttatcggtat
     1561 gttcatacag atgccgatca tgatcgcgta cgaaacagac cgtgccaccg gtgatggtat
     1621 agggctgccc attaaacaca tgaatacccg tgccatgttc gacaatcaca atttcatgaa
     1681 aatcatgatg atgttcagga aaatccgcct gcgggagccg gggttctatc gccacggacg
     1741 cgttaccaga cggaaaaaaa tccacactat gtaatacggt catactggcc tcctgatgtc
     1801 gtcaacacgg cgaaatagta atcacgaggt caggttctta ccttaaattt tcgacggaaa
     1861 accacgtaaa aaacgtcgat ttttcaagat acagcgtgaa ttttcaggaa atgcggtgag
     1921 catcacatca ccacaattca gcaaattgtg aacatcatca cgttcatctt tccctggttg
     1981 ccaatggccc attttcctgt cagtaacgag aaggtcgcga atacaggcgc tttttagact
     2041 ggtcgtaatg aaattcagca ggatggatct tacagtgagg cataattatg gctttcattc
     2101 caccaaccat cgacgacgtt agacattgct ctaacgcttt atctgtagac cccgccgaaa
     2161 ccgacgctgc ccgcgccatt gctgaacact actcaaagat atccaatcag gagtaccgca
     2221 tcacccaaga cgacctggat gatctcactg acacaatcga atatctcatg gccactaacc
     2281 agccagactc acaataagga tctccaggca tcaaataaaa cgaaaggctc agtcgaaaga
     2341 ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctctactaga gtcacactgg
     2401 ctcaccttcg ggtgggcctt tctgcgttta taggatccta actcgacgtg caggcttcct
     2461 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     2521 aggcggtaat caattcgacc cagctttctt gtacaaagtg gttgatcctt acagatcccg
     2581 gttatccaca gaatcagggg aggccttaga aatattttat ctgattaata agatgatctt
     2641 cttgagatcg ttttggtctg cgcgtaatct cttgctctga aaacgaaaaa accgccttgc
     2701 agggcggttt ttcgaaggtt ctctgagcta ccaactcttt gaaccgaggt aactggcttg
     2761 gaggagcgca gtcaccaaaa cttgtccttt cagtttagcc ttaaccggcg catgacttca
     2821 agactaactc ctctaaatca attaccagtg gctgctgcca gtggtgcttt tgcatgtctt
     2881 tccgggttgg actcaagacg atagttaccg gataaggcgc agcggtcgga ctgaacgggg
     2941 ggttcgtgca tacagtccag cttggagcga actgcctacc cggaactgag tgtcaggcgt
     3001 ggaatgagac aaacgcggcc ataacagcgg aatgacaccg gtaaaccgaa aggcaggaac
     3061 aggagagcgc acgagggagc cgccaggggg aaacgcctgg tatctttata gtcctgtcgg
     3121 gtttcgccac cactgatttg agcgtcagat ttcgtgatgc ttgtcagggg ggcggagcct
     3181 atggaaaaac ggctttgccg cggccctctc acttccctgt taagtatctt cctggcatct
     3241 tccaggaaat ctccgccccg ttcgtaagcc atttccgctc gccgcagtcg aacgaccgag
     3301 cgtagcgagt cagtgagcga ggaagcggaa tatatcctgt atcacatatt ctgctgacgc
     3361 accggtgcag ccttttttct cctgccacat gaagcacttc actgacaccc tcatcagtgc
     3421 caacatagta agccagtata cactccgcta gcgctgatgt ccggcggtgc gcatgccgtt
     3481 aagggatttt ggtcatgaac tagcttgatc gggcacgtaa gaggttccaa ctttcaccat
     3541 aatgaaataa gatcactacc gggcgtattt tttgagttat cgagattttc aggagctaag
     3601 gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat
     3661 cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt
     3721 cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg
     3781 gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaatttcg tatggcaatg
     3841 aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag
     3901 caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta
     3961 cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg
     4021 tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat
     4081 ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcaccatggg caaatattat
     4141 acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat
     4201 ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc
     4261 ggggcgtaat ttgatatcga gctcgcttgg actcctgttg atagatccag taatgacctc
     4321 agaactccat ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga
     4381 gaatccaagc ctgcagataa cttcgtatag catacattat acgaagttat ctcgaggacg
     4441 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     4501 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     4561 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     4621 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     4681 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     4741 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     4801 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     4861 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4921 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4981 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5041 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5101 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     5161 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     5221 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     5281 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     5341 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     5401 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     5461 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttgaa
     5521 ttgccattat caacaagttt gtacaaaaaa gcaggctccg aattggtatc acgaggcaga
     5581 atttcagata aaaaaaatcc ttagctttcg ctaaggatga tttctg
//
