LOCUS       sequence-444008-        5618 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  integrative plasmid for heterologous gene expression in S. pyogenes 
            SF370, integrates into the amyA locus via homologous recombination.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5618)
  AUTHORS   Lautenschlager N, Schmidt K, Schiffer C, Wulff TF, Hahnke K, 
            Finstermeier K, Mansour M, Elsholz AKW, Charpentier E
  TITLE     Expanding the genetic toolbox for the obligate human pathogen 
            Streptococcus pyogenes.
  JOURNAL   Front Bioeng Biotechnol. 2024 Jun 7;12:1395659. doi: 
            10.3389/fbioe.2024.1395659. eCollection 2024.
  PUBMED    38911550
REFERENCE   2  (bases 1 to 5618)
  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 
            Bioeng Biotechnol."; date: "2024-06-7"; volume: "12:1395659. doi"; 
            pages: " 10.3389/fbioe.2024.1395659. eCollection 2024"
COMMENT     Sequence Label: pEC2735 (pSpy0C2)
FEATURES             Location/Qualifiers
     source          1..5618
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(5..21)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5..21)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      2542..3130
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3031..3050
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3284..3301
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(4421..4455)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     primer_bind     4491..4507
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4617..4638
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4653..4658,4659..4676,4677..4683)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4691..4707
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             4725..4736
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4743..5420
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      5465..5536
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5552..5579
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
ORIGIN
        1 atgtgtcata gctgtttcct gcaaagaatg gtgatgacaa ttgtacaggt tatcaataaa
       61 gccattgata ctctcattct tatcccgtat ttattaactt ctcctagtat tagtaattat
      121 catggctgtc atggcgcatt aacggaataa agggtgtgct taaatcgggc cattttgcgt
      181 aataagaaaa aggattaatt atgagcgaat tgaattaata ataaggtaat agatttacat
      241 tagaaaatga aaggggattt tatgcgtgag aatgttacag tctatcccgg caatagttac
      301 ccttattatc aagataagaa agaaaaggat ttttcgctac gctcaaatcc tttaaaaaaa
      361 cacaaaagac cacatttttt aatgtggtct tttattcttc aactaaagca cccattagtt
      421 caacaaacga aaattggata aagtgggata tttttaaaat atatatttat gttacagtaa
      481 tattgacttt taaaaaagga ttgattataa tgaagaaagc agacaagtaa gcctcctaaa
      541 ttcactttag ataaaaattt aggaggcata tcaaatgaac tttaataaaa ttcactttag
      601 ataaaaattt aggaggcata tcaaatgaac tttaataaaa ttgatttaga caattggaag
      661 agaaaagaga tatttaatca ttatttgaac caacaaacga cttttagtat aaccacagaa
      721 attgatatta gtgttttata ccgaaacata aaacaagaag gatataaatt ttaccctgca
      781 tttattttct tagtgacaag ggtgataaac tcaaatacag cttttagaac tggttacaat
      841 agcgacggag agttaggtta ttgggataag ttagagccac tttatacaat ttttgatggt
      901 gtatctaaaa cattctctgg tatttggact cctgtaaaga atgacttcaa agagttttat
      961 gatttatacc tttctgatgt agagaaatat aatggttcgg ggaaattgtt tcccaaaaca
     1021 cctatacctg aaaatgcttt ttctctttct attattccat ggacttcatt tactgggttt
     1081 aacttaaata tcaataataa tagtaattac cttctaccca ttattacagc aggaaaattc
     1141 attaataaag gtaattcaat atatttaccg ctatctttac aggtacatca ttctgtttgt
     1201 gatggttatc atgcaggatt gtttatgaac tctattcagg aattgtcaga taggcctaat
     1261 gactggcttt tataatatga gataatgccg actgtacttt ttacagtcgg ttttctaatg
     1321 tcactaacct gccccgttag ttgaagaagg tttttatatt acagctccag atatgtatgg
     1381 tagagaacca aagacaacag aagcctaata aagccatgtg gctatctgtt atcccaggtt
     1441 tgggacaact atacaataaa caaatcgtta aagggggagt cttattagtt gttttcttcc
     1501 tagagctact agaaattgta gttctaggaa ttcctgcttt aacaggtctt tacagtcttg
     1561 gaagtgttcc aatgcaagat cactccttat ttatgttaat taagggggcc atgcaattaa
     1621 ttactctagt cttatttggt atttttcatt tggttgctat gagagatgct aaattagttg
     1681 cacatcaaat gaatgaaggc aagaaggttc cggtgacggc taaagagacc cttgaagcga
     1741 tttatgaaaa aggatttcct tacttgctca tcattccggc ttatttagca atggctttcg
     1801 ctattatttt tccagtttta gtgaccttgc tgattgcttt taccaattac gattttaggc
     1861 atattcctcc gtatcgttta ttggattggg tagggttgaa gaattttctt aatatcttag
     1921 aattaagcac cttcaaaaca gcatttacat ctgttttgtc atggacgctg atctggacat
     1981 tagcagcgac aagtcttcaa attgttattg gaattgcaac ggctgttatt gctaatcaat
     2041 cctttatcag aggaaaacgt ctctttggtg tgatattctt gttaccatgg gctgttccag
     2101 catttatctc tattatgagt tttggtaatt tctttaatga ttccattggg gccatgaatg
     2161 ttcaggttct tcctttcatc gagaaattcc ttcctttcgt tgacttcgga attgtgtcct
     2221 ggaagacaga ccccttctgg acaaaagtgg ctgttattat ggtgcaaggt tggctaggat
     2281 ttccctatat ctatattttg gtatcaggta ttttacaggc aattccagct gatttatacg
     2341 aagcagcaac cgttgatggt gcgacagcta actgtcagac caagtttact catatatact
     2401 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     2461 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     2521 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     2581 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     2641 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     2701 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     2761 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     2821 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     2881 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     2941 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     3001 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     3061 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     3121 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     3181 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     3241 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     3301 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     3361 atgcagctgg cacgacaggt ttcccgactg gaaagacctg aatcaagatt attctagacg
     3421 ctgtctttaa ccatgccagc agtgatgccg tggaatttca ggatgtcttg agatacggta
     3481 aagagtctaa gttttttgat tggtttatga ctcatgatga acacccatcc atggacctag
     3541 taaactatga aacctttgct ggatgtaact atatgcccaa atggaatacc tcaaatagag
     3601 acgtgcaaga ctatctgatt gagattgggc gatactggat taaagaattt tgtatcgatg
     3661 gttggcgcct agatgtttct gatgaagtat ctcatgattt ttggcgacgt ttccgtcagg
     3721 cggttaaagc ggaaaaagca gatgccattt taattgggga gaactggcat gatgcctatc
     3781 cttatttggc aggggatcag tatgatggga taatgaatta tgctttcacc aaagcctgtt
     3841 tggattactt tgcttttgaa accatgacat caaaacaatt ggctgagaaa ttaagcaaca
     3901 ttttgatgcg aaacacttgg caagtcaatc gcatgaactt gaatcttttg gattctcatg
     3961 atacgcatcg tttcttcact caagttaagg gttccaaaga taaattgtta gctgctttag
     4021 cattgctctt tatctttatg ggcattcctt gcttgtatta tgggacagaa gtggcaatgg
     4081 aagggggtta cgaccctgat tcgcgtcgcg gttttccttg ggatgatgct gagcgggaca
     4141 aggatttctt tagcaaagta aaagcgatta ttgctcttcg ccaacagcca gctcttcaag
     4201 aaggagctat tagcattgat tatgacgaag actgcctcat tctgaaacgc agcttaggtg
     4261 agcaaagcat ttcacttgtt cttaacaatg gtgagagtgc ttattctatt aaggaaggag
     4321 gacagatttt gacaagcaat ggggtaacta aaaacaaaaa acaattactc caggatggat
     4381 atattgtgtt tagtaactaa cagcgaacga atattggata ataaaaaacg cccggcggca
     4441 accgagcgtt ctgaattatg gggatgatgt taaggctatt ggtgtttatg gtaaaacgac
     4501 ggccagtcag cgagaccagc tgaattcaaa aaaaaatcta gatggccggc gattcattaa
     4561 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     4621 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     4681 ttgtgtggaa ttgtgagcgg ataacaattt cacacatact agagaaagag gagaaatact
     4741 agatggcttc ctccgaagac gttatcaaag agttcatgcg tttcaaagtt cgtatggaag
     4801 gttccgttaa cggtcacgag ttcgaaatcg aaggtgaagg tgaaggtcgt ccgtacgaag
     4861 gtacccagac cgctaaactg aaagttacca aaggtggtcc gctgccgttc gcttgggaca
     4921 tcctgtcccc gcagttccag tacggttcca aagcttacgt taaacacccg gctgacatcc
     4981 cggactacct gaaactgtcc ttcccggaag gtttcaaatg ggaacgtgtt atgaacttcg
     5041 aagacggtgg tgttgttacc gttacccagg actcctccct gcaagacggt gagttcatct
     5101 acaaagttaa actgcgtggt accaacttcc cgtccgatgg tccggttatg cagaaaaaaa
     5161 ccatgggttg ggaagcttcc accgaacgta tgtacccgga agacggtgct ctgaaaggtg
     5221 aaatcaaaat gcgtctgaaa ctgaaagacg gtggtcacta cgacgctgaa gttaaaacca
     5281 cctacatggc taaaaaaccg gttcagctgc cgggtgctta caaaaccgac atcaaactgg
     5341 acatcacctc ccacaacgaa gactacacca tcgttgaaca gtacgaacgt gctgaaggtc
     5401 gtcactccac cggtgcttaa taacgctgat agtgctagtg tagatcgcta ctagagccag
     5461 gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt
     5521 gtcggtgaac gctctctact agagtcacac tggctcacct tcgggtgggc ctttctgcgt
     5581 ttataaccgg ttaatactag tactgcagtg aaggtctc
//
