LOCUS       sequence-329699-        4658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SiMPl plasmid pair (pSiMPls) for use with 
            spectinomycin/streptomycin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4658)
  AUTHORS   Palanisamy N, Ballestin JB, Di Ventura B
  TITLE     Expanding the SiMPl Plasmid Toolbox for Use with 
            Spectinomycin/Streptomycin.
  JOURNAL   ACS Omega. 2021 May 21;6(22):14148-14153. doi: 
            10.1021/acsomega.1c00649. eCollection 2021 Jun 8.
  PUBMED    34124437
REFERENCE   2  (bases 1 to 4658)
  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/acsomega.1c00649"; journalName: "ACS Omega"; date: 
            "2021-05-21- 21"; volume: "6"; issue: "22"; pages: "14148-14153"
COMMENT     Sequence Label: pSiMPls_N + pSiMPls_C
FEATURES             Location/Qualifiers
     source          1..4658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(193..198,199..215,216..222)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    230..246
                     /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)."
     primer_bind     235..257
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             276..983
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTDI
                     ALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAVA
                     KYSNLGGGMDELYK"
     primer_bind     complement(1025..1042)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1195..1281
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1373..1400
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1419..1510
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2359..2947
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2848..2867
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3101..3118
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3133..3273
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3199..3218)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3359..3381
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        3459..3536
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3537..4619
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3556..3575)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc
       61 ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc
      121 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
      181 tgaaatgagc tgttgacaat taatcatccg gctcgtataa tgtgtggaat tgtgagcgga
      241 taacaatttc acacaggaaa cagaccatgg aattcgtgtc taagggcgaa gagctgatca
      301 aggaaaatat gcgtatgaag gtggtcatgg aaggttcggt caacggccac caattcaaat
      361 gcacaggtga aggagaaggc agaccgtacg agggagtgca aaccatgagg atcaaagtca
      421 tcgagggagg acccctgcca tttgcctttg acattcttgc cacgtcgttc atgtatggca
      481 gccgtacctt tatcaagtac ccggccgaca tccctgattt ctttaaacag tcctttcctg
      541 agggttttac ttgggaaaga gttacgagat acgaagatgg tggagtcgtc accgtcacgc
      601 aggacaccag ccttgaggat ggcgagctcg tctacaacgt caaggtcaga ggggtaaact
      661 ttccctccaa tggtcccgtg atgcagaaga agaccaaggg ttgggagcct aatacagaga
      721 tgatgtatcc agcagatggt ggtctgagag gatacactga catcgcactg aaagttgatg
      781 gtggtggcca tctgcactgc aacttcgtga caacttacag gtcaaaaaag accgtcggga
      841 acatcaagat gcccggtgtc catgccgttg atcaccgcct ggaaaggatc gaggagagtg
      901 acaatgaaac ctacgtagtg caaagagaag tggcagttgc caaatacagc aaccttggtg
      961 gtggcatgga cgagctgtac aagtaaaagc ttggctgttt tggcggatga gagaagattt
     1021 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     1081 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     1141 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     1201 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     1261 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     1321 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     1381 atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca
     1441 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     1501 aaggaagagt atgatgctga aaaaaatcct gaaaatcgaa gagctggatg aacgtgaact
     1561 gatcgatatt gaggtgtccg gtaaccacct gttttacgct aacgatattc tgacccacaa
     1621 ctcccctgga gagagcgaga ttctccgcgc tgtagaagtc accattgttg tgcacgacga
     1681 catcattccg tggcgttatc cagctaagcg cgaactgcaa tttggagaat ggcagcgcaa
     1741 tgacattctt gcaggtatct tcgagccagc cacgatcgac attgatctgg ctatcttgct
     1801 gacaaaagca agagaacata gcgttgcctt ggtaggtcca gcggcggagg aactctttga
     1861 tccggttcct gaacaggatc tatttgaggc gctaaatgaa accttaacgc tatggaactc
     1921 gccgcccgac tgggctggcg atgagcgaaa tgtagtgctt acgttgtccc gcatttggta
     1981 cagcgcagta accggcaaaa tcgcgccgaa ggatgtcgct gccgactggg caatggagcg
     2041 cctgccggcc cagtatcagc ccgtcatact tgaagctaga caggcttatc ttggacaaga
     2101 agaagatcgc ttggcctcgc gcgcagatca gttggaagaa tttgtccact acgtgaaagg
     2161 cgagatcacc aaggtagtcg gcaaataact gtcagaccaa gtttactcat atatacttta
     2221 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     2281 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2341 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2401 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2461 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     2521 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2581 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2641 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2701 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2761 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2821 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2881 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     2941 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     3001 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     3061 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     3121 agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3181 catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     3241 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
     3301 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     3361 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagcagatc
     3421 aattcgcgcg cgaaggcgaa gcggcatgca tttacgttga caccatcgaa tggtgcaaaa
     3481 cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga
     3541 aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc
     3601 gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga
     3661 tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt
     3721 tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg
     3781 cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa
     3841 gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc
     3901 tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta
     3961 atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt attattttct
     4021 cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa
     4081 tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc
     4141 ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg
     4201 ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga
     4261 tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc
     4321 tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt
     4381 atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg
     4441 accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct
     4501 cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt
     4561 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     4621 cgcaacgcaa ttaatgtaag ttagcgcgaa ttgatctg
//
