LOCUS       sequence-395257-        6182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Arabinose induced expression of wildtype repL gene, which triggers 
            DNA replication in cis and in trans, weak level of constitutive GFP 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6182)
  AUTHORS   Huan YW, Brown R, Wang B
  TITLE     An adenine/thymidine-rich region is integral to RepL-mediated DNA 
            replication.
  JOURNAL   Front Microbiol. 2023 Feb 9;14:1095671. doi: 
            10.3389/fmicb.2023.1095671. eCollection 2023.
  PUBMED    36846746
REFERENCE   2  (bases 1 to 6182)
  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"; journalName: "Front 
            Microbiol."; date: "2023-02-9"; volume: "14:1095671. doi"; pages: " 
            10.3389/fmicb.2023.1095671. eCollection 2023"
COMMENT     Sequence Label: 4A3.PBAD-repL(WT).J23115-gfp
FEATURES             Location/Qualifiers
     source          1..6182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(226..330)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(365..392)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(408..479)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(502..1215)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(542..562)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     1157..1185
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     RBS             1222..1233
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(1245..1279)
                     /label=BBa_J23115
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.15"
     CDS             complement(1331..2209)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        2236..2520
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     2443..2462
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             complement(3711..4661)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      4709..4931
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(5423..5517)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(5547..6182,1..156,157..225))
                     /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     join(6170..6182,1..7)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
       61 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
      121 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
      181 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
      241 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
      301 ttagaaaaat aaacaaatag gggttccgcg cacatttccc atggtgccac ctgacgtcta
      361 agaacgcaga aaggcccacc cgaaggtgag ccagtgtgac tctagtagag agcgttcacc
      421 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga
      481 tgcctggctc tagtattatt atttgtatag ttcatccatg ccatgtgtaa tcccagcagc
      541 tgttacaaac tcaagaagga ccatgtggtc tctcttttcg ttgggatctt tcgaaagggc
      601 agattgtgtg gacaggtaat ggttgtctgg taaaaggaca gggccatcgc caattggagt
      661 attttgttga taatggtctg ctagttgaac gcttccatct tcaatgttgt gtctaatttt
      721 gaagttaact ttgattccat tcttttgttt gtctgccatg atgtatacat tgtgtgagtt
      781 atagttgtat tccaatttgt gtccaagaat gtttccatct tctttaaaat caataccttt
      841 taactcgatt ctattaacaa gggtatcacc ttcaaacttg acttcagcac gtgtcttgta
      901 gttcccgtca tctttgaaaa atatagttct ttcctgtaca taaccttcgg gcatggcact
      961 cttgaaaaag tcatgctgtt tcatatgatc tgggtatctc gcaaagcatt gaacaccata
     1021 accgaaagta gtgacaagtg ttggccatgg aacaggtagt tttccagtag tgcaaataaa
     1081 tttaagggta agttttccgt atgttgcatc accttcaccc tctccactga cagaaaattt
     1141 gtgcccatta acatcaccat ctaattcaac aagaattggg acaactccag tgaaaagttc
     1201 ttctccttta cgcatctagt atttctcctc tttaatctct agtagctagc attgtaccaa
     1261 gggctgagct agctataaat tttgttatca ataaaaaagg ccgcgatttg cggccttatt
     1321 gttcgtctta ttatgacaac ttgacggcta catcattcac tttttcttca caaccggcac
     1381 gaaactcgct cgggctggcc ccggtgcatt ttttaaatac tcgcgagaaa tagagttgat
     1441 cgtcaaaacc aacattgcga ccgacggtgg cgataggcat ccgggtagtg ctcaaaagca
     1501 gcttcgcctg actaatgcgt tggtcctcgc gccagcttaa gacgctaatc cctaactgct
     1561 ggcggaaaag atgtgacaga cgcgacggcg acaagcaaac atgctgtgcg acgctggcga
     1621 tatcaaaatt gctgtctgcc aggtgatcgc tgatgtactg acaagcctcg cgtacccgat
     1681 tatccatcgg tggatggagc gactcgttaa tcgcttccat gcgccgcagt aacaattgct
     1741 caagcagatt tatcgccagc agctccgaat agcgcccttc cccttgcccg gcgttaatga
     1801 tttgcccaaa caggtcgctg aaatgcggct ggtgcgcttc atccgggcga aagaaacccg
     1861 tattggcaaa tattgacggc cagttaagcc attcatgcca gtaggcgcgc ggacgaaagt
     1921 aaacccactg gtgataccat tcgcgagcct ccggatgacg accgtagtga tgaatctctc
     1981 ctggcgggaa cagcaaaata tcacccggtc ggcagacaaa ttctcgtccc tgatttttca
     2041 ccaccccctg accgcgaatg gtgagattga gaatataacc tttcattccc agcggtcggt
     2101 cgataaaaaa atcgagataa ccgttggcct caatcggcgt taaacccgcc accagatggg
     2161 cgttaaacga gtatcccggc agcaggggat cattttgcgc ttcagccata cttttcatac
     2221 tcccaccatt cagagaagaa accaattgtc catattgcat cagacattgc cgtcactgcg
     2281 tcttttactg gctcttctcg ctaacccaac cggtaacccc gcttattaaa agcattctgt
     2341 aacaaagcgg gaccaaagcc atgacaaaaa cgcgtaacaa aagtgtctat aatcacggca
     2401 gaaaagtcca cattgattat ttgcacggcg tcacactttg ctatgccata gcatttttat
     2461 ccataagatt agcggatcct acctgacgct ttttatcgca actctctact gtttctccat
     2521 gtacctgtcg cggcaacgcg ctaacagacg tagtaagaac caccagcatt gtaatgctgg
     2581 ctaaagtcac tttcctgagc tgtataacga tgagcgattt tactttttct ggctatgaat
     2641 tggcctgctt tgtaacacac tccggtctat cccgtagcgc cgggcatatc ctgtcgcaat
     2701 gtgcaaatct cgcggcaaca accagtgaat acttcattca caagcctcac cgcctgatcg
     2761 cggcagaaac tggttatagc caatcaaccg tcgttcgtgc attccgtgaa gctgtaaaca
     2821 aaggaattct gtctgtagag attgttatcg gcgatcaccg tgaacgtcgc gctaacctgt
     2881 accggtttac accatccttt ttggccttcg cacaacaagc caaaaatgcg ctgatagaaa
     2941 gcaaattaaa gatctcttca gcggcaacca aggttaaagc tgttctcgct aagacattgg
     3001 ctttatttaa ttttttatcc acacccccat gtcaaaatga taccccctcc ccctgtcagg
     3061 atgacgtggc aataaagaat aagaagtcac aagttaaaaa aacaaaaaga tcagtttccg
     3121 gcggtgccgg aacaaccagc ctcaaaaaat tgacttcatg gatcgctaag gcaaaagcaa
     3181 aggctgacaa tctgcggtta tccaaaaaac gcactcaaaa acatgagttc aagcagaaag
     3241 tagaggcggc tgcgcggaaa tatgcttacc tgaagaacaa gcgttcgcct gatattggcg
     3301 ggatatcaaa cttcgataac ctaccgcatt gcatgacggt aaacgaagct cttaatgcgg
     3361 ttttagccaa aaataaagat aacgaacaat ggggtatacc ggcaggattc agagggtaat
     3421 aactcggtac caaattccag aaaagaggcc tcccgaaagg ggggcctttt ttcgttttgg
     3481 tccgctcact caaaggcggt aatctcgagg ttacattgtc gatctgttca tggtgaacag
     3541 ctttgaatgc accaaaaact cgtaaaagct ctgatgtatc tatctttttt acaccgtttt
     3601 catctgtgca tatggacagt tttccctttg atatgtaacg gtgaacagtt gttctacttt
     3661 tgtttgttag tcttgatgct tcactgatag atacaagagc cataagaacc tcagatcctt
     3721 ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag ccatgagaac
     3781 gaaccattga gatcatactt actttgcatg tcactcaaaa attttgcctc aaaactggtg
     3841 agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt atgtaggtag
     3901 gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg gttgttctca
     3961 agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa cgtatcagtc
     4021 gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa atctttactt
     4081 attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt ttcaagcatt
     4141 aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt tttcttttgt
     4201 gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc aaaagactta
     4261 acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg caatatctct
     4321 tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt ccactggaaa
     4381 atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat cagctctctg
     4441 gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat ctgagcgtat
     4501 tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat cgtggggttg
     4561 agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc atagcgacta
     4621 atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa ttggtctagg
     4681 tgattttaat cactatacca attgagatgg gctagtcaat gataattaca tgtccttttc
     4741 ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact tgtaaattct
     4801 gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta tattcaagtg
     4861 gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga tcccagccct
     4921 gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc gcaaacgctg
     4981 tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac cctcgcaagc
     5041 tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga gtcgctgtct
     5101 ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg gtaaatggca
     5161 ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag aaaagcccgt
     5221 cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct gactttttgc
     5281 tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta tcccgtgaca
     5341 ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca ggcttacccg
     5401 tcttactgtc cctagtgctt ggattctcac caataaaaaa cgcccggcgg caaccgagcg
     5461 ttctgaacaa atccagatgg agttctgagg tcattactgg atctatcaac aggagtccaa
     5521 gcgagctcgt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     5581 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5641 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     5701 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5761 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     5821 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     5881 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     5941 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6001 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6061 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6121 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6181 cc
//
