LOCUS       sequence-415309-        4709 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for hlpA-mTurquoise2 amplification associated with 
            erythromycin resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4709)
  AUTHORS   Mignolet J, Staub J, van der Meer JR, Veening J-W, Libante V
  TITLE     A Modular Genetic Toolbox for Precise Gene Regulation and 
            Multi-Color Imaging in Streptococci
  JOURNAL   bioRxiv 2025.05.10.653270
REFERENCE   2  (bases 1 to 4709)
  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: "bioRxiv 
            2025.05.10.653270"
COMMENT     Sequence Label: pJethlpAmTurquoise2Ery
FEATURES             Location/Qualifiers
     source          1..4709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     305..324
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        305..323
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(685..687,688..1401)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(685..687,688..1401)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2010)"
                     /label=mTurquoise
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(727..748)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(988..1007)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1335..1356
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(join(2541..2547,2548..2565,2566..2571))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    2586..2607
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2727..2744)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2898..3486)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2978..2997)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3657..4448,4449..4517))
                     /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     4280..4299
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4518..4622)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4690..4708
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcccctgcag ccgaattata ttatttttgc caaataattt ttaacaaaag ctctgaagtc
       61 ttcttcattt aaattcttag atgatacttc atctggaaaa ttgtcccaat tagtagcatc
      121 acgctgtgag taagttctaa accatttttt tattgttgta ttatctctaa tcttactact
      181 cgatgagttt tcggtattat ctctattttt aacttggagc aggttccatt cattgttttt
      241 ttcatcatag tgaataaaat caactgcttt aacacttgtg cctgaacacc atatccatcc
      301 ggcgtaatac gactcactat agggagagcg gccgccagat cttccggatg gctcgagttt
      361 ttcagcaaga tcatggcaaa caaacaagat ttgatcgcta aagtagcaga agctacagaa
      421 ttgactaaga aagactcagc agcagcagtt gaagctgtat ttgtagcagt agctgactat
      481 cttgcagctg gtgaaaaagt tcaattgatc ggttttggta actttgaagt tcgtgagcgt
      541 gcagaacgta aaggtcgcaa cccacaaact ggtaaagaaa tgacaattgc agcttctaaa
      601 gtaccagcat tcaaagctgg taaagctctt aaagacgctg ttaaaggatc cggatctggt
      661 ggagaagctg aagctaaagg atcagtgagc aagggcgagg agctgttcac cggggtggtg
      721 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
      781 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
      841 ctgcccgtgc cctggcccac cctcgtgacc accctgtcct ggggcgtgca gtgcttcgcc
      901 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
      961 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1021 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1081 gacggcaaca tcctggggca caagctggag tacaactact tcagcgacaa cgtctatatc
     1141 accgccgaca agcagaagaa cggcatcaag gccaacttca agatccgcca caacatcgag
     1201 gacggcggcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1261 gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac
     1321 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1381 atggacgagc tgtacaagta ataagaattc caaacttaag agtgtgttga tagtgcagta
     1441 tcttaaaatt ttgtataata ggaattgaag ttaaattaga tgctaaaaat ttgtaattaa
     1501 gaaggagtga ttacatgaac aaaaatataa aatattctca aaacttttta acgagtgaaa
     1561 aagtactcaa ccaaataata aaacaattga atttaaaaga aaccgatacc gtttacgaaa
     1621 ttggaacagg taaagggcat ttaacgacga aactggctaa aataagtaaa caggtaacgt
     1681 ctattgaatt agacagtcat ctattcaact tatcgtcaga aaaattaaaa ctgaatactc
     1741 gtgtcacttt aattcaccaa gatattctac agtttcaatt ccctaacaaa cagaggtata
     1801 aaattgttgg gagtattcct taccatttaa gcacacaaat tattaaaaaa gtggtttttg
     1861 aaagccatgc gtctgacatc tatctgattg ttgaagaagg attctacaag cgtaccttgg
     1921 atattcaccg aacactaggg ttgctcttgc acactcaagt ctcgattcag caattgctta
     1981 agctgccagc ggaatgcttt catcctaaac caaaagtaaa cagtgtctta ataaaactta
     2041 cccgccatac cacagatgtt ccagataaat attggaagct atatacgtac tttgtttcaa
     2101 aatgggtcaa tcgagaatat cgtcaactgt ttactaaaaa tcagtttcat caagcaatga
     2161 aacacgccaa agtaaacaat ttaagtaccg ttacttatga gcaagtattg tctattttta
     2221 atagttatct attatttaac gggaggaaat aattctatga gtcgcttttg taaatttgga
     2281 aagttacacg ttactaaagg gaatgtagat aaattattag gtatactact gacagcttcc
     2341 aaggagctaa agaggtccaa gcttcaggaa actgagacag gaattttatt aaaaatttaa
     2401 attttgaaga aagttcaggg ttaatagcat ccattttttg ctttgcaagt tcctcagcat
     2461 tcttaacaaa agacgtctct tttgacatgt ttaaagttta aacctcctgt gtgaaattat
     2521 tatccgctca taattccaca cattatacga gccggaagca taaagtgtaa agcctggggt
     2581 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgccaat tgctttccag
     2641 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     2701 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     2761 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     2821 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     2881 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     2941 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     3001 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     3061 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     3121 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     3181 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     3241 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     3301 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     3361 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     3421 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     3481 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     3541 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     3601 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     3661 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     3721 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     3781 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     3841 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     3901 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     3961 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     4021 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     4081 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     4141 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     4201 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     4261 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     4321 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     4381 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     4441 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     4501 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     4561 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     4621 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     4681 acctataaaa ataggcgtat cacgaggcc
//
