LOCUS       sequence-324733-        4659 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 4659)
  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 4659)
  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..4659
                     /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        1420..1511
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2360..2948
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2849..2868
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3102..3119
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3134..3274
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3200..3219)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3360..3382
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        3460..3537
                     /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             3538..4620
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3557..3576)
                     /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 ttgtttattt ttctaaatac
     1441 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1501 aaaggaagag tatgatgctg aaaaaaatcc tgaaaatcga agagctggat gaacgtgaac
     1561 tgatcgatat tgaggtgtcc ggtaaccacc tgttttacgc taacgatatt ctgacccaca
     1621 actcccctgg agagagcgag attctccgcg ctgtagaagt caccattgtt gtgcacgacg
     1681 acatcattcc gtggcgttat ccagctaagc gcgaactgca atttggagaa tggcagcgca
     1741 atgacattct tgcaggtatc ttcgagccag ccacgatcga cattgatctg gctatcttgc
     1801 tgacaaaagc aagagaacat agcgttgcct tggtaggtcc agcggcggag gaactctttg
     1861 atccggttcc tgaacaggat ctatttgagg cgctaaatga aaccttaacg ctatggaact
     1921 cgccgcccga ctgggctggc gatgagcgaa atgtagtgct tacgttgtcc cgcatttggt
     1981 acagcgcagt aaccggcaaa atcgcgccga aggatgtcgc tgccgactgg gcaatggagc
     2041 gcctgccggc ccagtatcag cccgtcatac ttgaagctag acaggcttat cttggacaag
     2101 aagaagatcg cttggcctcg cgcgcagatc agttggaaga atttgtccac tacgtgaaag
     2161 gcgagatcac caaggtagtc ggcaaataac tgtcagacca agtttactca tatatacttt
     2221 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     2281 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2341 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2401 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2461 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     2521 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     2581 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     2641 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     2701 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     2761 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     2821 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     2881 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     2941 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3001 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     3061 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     3121 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3181 gcatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     3241 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     3301 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     3361 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagcagat
     3421 caattcgcgc gcgaaggcga agcggcatgc atttacgttg acaccatcga atggtgcaaa
     3481 acctttcgcg gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg
     3541 aaaccagtaa cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc
     3601 cgcgtggtga accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg
     3661 atggcggagc tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg
     3721 ttgctgattg gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg
     3781 gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga
     3841 agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg
     3901 ctgatcatta actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact
     3961 aatgttccgg cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc
     4021 tcccatgaag acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa
     4081 atcgcgctgt tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg
     4141 cataaatatc tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt
     4201 gccatgtccg gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg
     4261 atgctggttg ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg
     4321 ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg ataccgaaga cagctcatgt
     4381 tatatcccgc cgtcaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg
     4441 gaccgcttgc tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgcccgtc
     4501 tcactggtga aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg
     4561 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     4621 gcgcaacgca attaatgtga gttagcgcga attgatctg
//
