LOCUS       sequence-272560-        4340 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Maintenance of 2 plasmids with a single antibiotic in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4340)
  AUTHORS   Palanisamy N, Degen A, Morath A, Ballestin Ballestin J, Juraske C, 
            Ozturk MA, Sprenger GA, Youn JW, Schamel WW, Di Ventura B
  TITLE     Split intein-mediated selection of cells containing two plasmids 
            using a single antibiotic.
  JOURNAL   Nat Commun. 2019 Oct 31;10(1):4967. doi: 10.1038/s41467-019-12911-1.
  PUBMED    31672972
REFERENCE   2  (bases 1 to 4340)
  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: "Nat 
            Commun."; date: "2019-10-31"; volume: "10(1):4967. doi"; pages: " 
            10.1038/s41467-019-12911-1"
COMMENT     Sequence Label: pSiMPlc_N + pSiMPlc_C
FEATURES             Location/Qualifiers
     source          1..4340
                     /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      2041..2629
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2530..2549
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2783..2800
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2815..2955
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2881..2900)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3041..3063
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        3141..3218
                     /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             3219..4301
                     /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(3238..3257)
                     /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 ctcttcttct aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga
     1681 tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta
     1741 tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga
     1801 tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg
     1861 cggggcgtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     1921 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     1981 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2041 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2101 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2161 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     2221 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2281 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2341 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2401 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     2461 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     2521 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2581 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2641 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     2701 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     2761 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     2821 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt gcactctcag
     2881 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     2941 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3001 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3061 aggttttcac cgtcatcacc gaaacgcgcg aggcagcaga tcaattcgcg cgcgaaggcg
     3121 aagcggcatg catttacgtt gacaccatcg aatggtgcaa aacctttcgc ggtatggcat
     3181 gatagcgccc ggaagagagt caattcaggg tggtgaatgt gaaaccagta acgttatacg
     3241 atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca
     3301 gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag ctgaattaca
     3361 ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt ggcgttgcca
     3421 cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg
     3481 atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta
     3541 aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc
     3601 tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg gcgttatttc
     3661 ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgaa gacggtacgc
     3721 gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc
     3781 cattaagttc tgtctcggcg cgtctgcgtc tggctggctg gcataaatat ctcactcgca
     3841 atcaaattca gccgatagcg gaacgggaag gcgactggag tgccatgtcc ggttttcaac
     3901 aaaccatgca aatgctgaat gagggcatcg ttcccactgc gatgctggtt gccaacgatc
     3961 agatggcgct gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata
     4021 tctcggtagt gggatacgac gataccgaag acagctcatg ttatatcccg ccgttaacca
     4081 ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct
     4141 ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa
     4201 ccaccctggc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     4261 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgta
     4321 agttagcgcg aattgatctg
//
