LOCUS       sequence-409938-        4945 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for hlpA-mTurquoise2 amplification associated with 
            chloramphenicol resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4945)
  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 4945)
  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: pJethlpAmTurquoise2Cm
FEATURES             Location/Qualifiers
     source          1..4945
                     /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(2777..2783,2784..2801,2802..2807))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    2822..2843
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2963..2980)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3134..3722)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3214..3233)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3893..4684,4685..4753))
                     /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     4516..4535
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4754..4858)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4926..4944
                     /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 cgtattggtg gaataacacg aattaataaa
     1441 ggatatattt gtagtagcaa gtgtatttgt tttatattct atgaccctat ttgatttaca
     1501 tgattctata tggtataatg gacttgctta taaatataga tgcttggata gcaggaggtt
     1561 gcaagatgat gatggcagtt aaattagcct tctaaaatta aataataaaa ttttaggagg
     1621 ctaatttatg aaatttaatt tgatagatat agaggattgg aatagaaagc catactttga
     1681 gcattattta aatgcggtaa ggtgcactta cagtatgact gcaaatttag agataactag
     1741 tttactgcgt gaaattaaac ttaagggcct gaaactgtac cctacgctta tttatatcat
     1801 cacaactgta gttaatcgtc acaaggagtt ccgcaccagt tttgatcaaa acggtaagtt
     1861 aggatactgg gatagtatga acccaagtta tactatcttt cataaggata acgaaacttt
     1921 ttcaagtatt tggacagagt atgacgagaa cttcccacgt ttctactata attaccttga
     1981 ggatatcaga aactatagcg acgttatgaa tttcatgcct aaggcaggtc aacctgctaa
     2041 cacaattaat gtgtccagca ttccttgggt gaattttacc ggattcaact tgaatatata
     2101 taatgatgca acatatctaa tccctatttt tactatgggt aagtattttc agcaggataa
     2161 taaaatttta ttacctatgt ctgtacaggt gcatcatgcg gtttgcgacg gttaccatac
     2221 cagcagattt tttaatgagg tacaggaatt agtctcaaat tttgagacat ggttaggaga
     2281 aaaataatat taaaatgctt gtgctcggga gttataaaca aagatcgaaa atattgttaa
     2341 gatcgtgagg aagcaaagcg accgaagaag tatatgttcc atctatgcaa ggacttcccg
     2401 agaaggcgag tgacagcgaa gaccgattgg aaggagtctg agctaagtgg agtaatatat
     2461 tcattcatta tagaaccatc atttcaaagg attagagaaa tctaatcctt tttctaggta
     2521 tttttatcct aagaatttaa tatgacaatc acgagtttaa tttttatttt ggtgctattt
     2581 atcctaagca gagcaagctt caggaaactg agacaggaat tttattaaaa atttaaattt
     2641 tgaagaaagt tcagggttaa tagcatccat tttttgcttt gcaagttcct cagcattctt
     2701 aacaaaagac gtctcttttg acatgtttaa agtttaaacc tcctgtgtga aattattatc
     2761 cgctcataat tccacacatt atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     2821 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gccaattgct ttccagtcgg
     2881 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     2941 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3001 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3061 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3121 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3181 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3241 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3301 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     3361 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     3421 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     3481 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     3541 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     3601 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3661 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3721 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3781 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3841 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     3901 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     3961 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4021 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4081 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4141 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4201 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4261 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4321 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4381 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     4441 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     4501 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     4561 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4621 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4681 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4741 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     4801 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     4861 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     4921 ataaaaatag gcgtatcacg aggcc
//
