LOCUS       sequence-101265-        5113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Positive control duet plasmid for SepOTS-dependent split mCherry 
            assembly assay (i.e., N-mCherry is fused to the peptide ME(pS)VD (pS
            = phosphoserine); C-mCherry is fused to TRAP(RK)).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5113)
  AUTHORS   Sawyer N, Gassaway BM, Haimovich AD, Isaacs FJ, Rinehart J, Regan L
  TITLE     Designed Phosphoprotein Recognition in Escherichia coli.
  JOURNAL   ACS Chem Biol. 2014 Nov 21;9(11):2502-7. doi: 10.1021/cb500658w. 
            Epub 2014 Oct 6.
  PUBMED    25272187
REFERENCE   2  (bases 1 to 5113)
  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/cb500658w";
            journalName: "ACS Chem Biol"; date: "2014-11-21- 21"; volume: "9"; 
            issue: "11"; pages: "2502-7"
COMMENT     Sequence Label: pNAS1B split mCherry N-ME(pS)VD/TRAP(RK)-C
FEATURES             Location/Qualifiers
     source          1..5113
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(7..25,26..31,32..48,49..54,55..80)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    7..25
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    32..50
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             90..101
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(259..277)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     527..546
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      669..755
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(909..926)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1043..1588)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1123..1142)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1736..2614)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        2641..2925
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     2848..2867
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     primer_bind     complement(3645..3662)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3815..3901
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3993..4020
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4050..4113
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4114..4182,4183..4974)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4332..4351)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 agatcttccc tatcagtgat agagattgac atccctatca gtgatagaga tactgagcac
       61 atcagcagga cgcactgacc gaattcatta aagaggagaa aggtaccatg gcatccgtga
      121 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
      181 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
      241 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
      301 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
      361 acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
      421 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
      481 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga
      541 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggtggctctg
      601 gctctgggtc gactggtatg gaataggtgg attaatgaaa gcttcatggt acgcgtgctg
      661 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
      721 gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gccctcctag gatcagcgct
      781 agcggagtgt atactggctt actatgttgg cactgatgag ggtgtcagtg aagtgcttca
      841 tgtggcagga gaaaaaaggc tgcaccggtg cgtcagcaga atatgtgata caggatatat
      901 tccgcttcct cgctcactga ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg
      961 gcttacgaac ggggcggaga tttcctggaa gatgccagga agatacttaa cagggaagtg
     1021 agagggccgc ggcaaagccg tttttccata ggctccgccc ccctgacaag catcacgaaa
     1081 tctgacgctc aaatcagtgg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1141 cccctggcgg ctccctcgtg cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc
     1201 cgctgttatg gccgcgtttg tctcattcca cgcctgacac tcagttccgg gtaggcagtt
     1261 cgctccaagc tggactgtat gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc
     1321 ggtaactatc gtcttgagtc caacccggaa agacatgcaa aagcaccact ggcagcagcc
     1381 actggtaatt gatttagagg agttagtctt gaagtcatgc gccggttaag gctaaactga
     1441 aaggacaagt tttggtgact gcgctcctcc aagccagtta cctcggttca aagagttggt
     1501 agctcagaga accttcgaaa aaccgccctg caaggcggtt ttttcgtttt cagagcaaga
     1561 gattacgcgc agaccaaaac gatctcaaga agatcatctt attaactgca gagatcaatt
     1621 cgcgcgcgaa ggcgaagcgg catgctgcat aatgtgcctg tcaaatggac gaagcaggga
     1681 ttctgcaaac cctatgctac tccgtcaagc cgtcaattgt ctgattcgtt accaattatg
     1741 acaacttgac ggctacatca ttcacttttt cttcacaacc ggcacggaac tcgctcgggc
     1801 tggccccggt gcatttttta aatacccgcg agaaatagag ttgatcgtca aaaccaacat
     1861 tgcgaccgac ggtggcgata ggcatccggg tggtgctcaa aagcagcttc gcctggctga
     1921 tacgttggtc ctcgcgccag cttaagacgc taatccctaa ctgctggcgg aaaagatgtg
     1981 acagacgcga cggcgacaag caaacatgct gtgcgacgct ggcgatatca aaattgctgt
     2041 ctgccaggtg atcgctgatg tactgacaag cctcgcgtac ccgattatcc atcggtggat
     2101 ggagcgactc gttaatcgct tccatgcgcc gcagtaacaa ttgctcaagc agatttatcg
     2161 ccagcagctc cgaatagcgc ccttcccctt gcccggcgtt aatgatttgc ccaaacaggt
     2221 cgctgaaatg cggctggtgc gcttcatccg ggcgaaagaa ccccgtattg gcaaatattg
     2281 acggccagtt aagccattca tgccagtagg cgcgcggacg aaagtaaacc cactggtgat
     2341 accattcgcg agcctccgga tgacgaccgt agtgatgaat ctctcctggc gggaacagca
     2401 aaatatcacc cggtcggcaa acaaattctc gtccctgatt tttcaccacc ccctgaccgc
     2461 gaatggtgag attgagaata taacctttca ttcccagcgg tcggtcgata aaaaaatcga
     2521 gataaccgtt ggcctcaatc ggcgttaaac ccgccaccag atgggcatta aacgagtatc
     2581 ccggcagcag gggatcattt tgcgcttcag ccatactttt catactcccg ccattcagag
     2641 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
     2701 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
     2761 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
     2821 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
     2881 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
     2941 taacaggagg aatgacgtca tggctaagca ggcactgaaa gaaaaagagc tggggaacga
     3001 tgcctacaag aagaaagact ttgacacagc cttgaagcat tacgacaaag ccaaggagct
     3061 ggaccccact aacatgactt acattaccaa tcaagcagcg gtatactttg aaaagggcga
     3121 ctacaataag tgccgggagc tttgtgagaa ggccattgaa gtggggagag aaaaccgaga
     3181 agactatcga cagattgcca aagcatatgc tcgaattggc aactcctact tcaaagaaga
     3241 aaagtacaag gatgccatcc atttctataa caagtctctg gcagaacacc gacgcccaaa
     3301 agtgctcaag aaatgccagc aggcagagaa aatcctgaag gagcaagcgg ccgcaggtgg
     3361 aagcggtggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca
     3421 ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc
     3481 ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga
     3541 acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagtg
     3601 ataagagctc ttggctgttt tggcggatga gagaagattt tcagcctgat acagattaaa
     3661 tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag cgcggtggtc
     3721 ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg
     3781 tctccccatg cgagagtagg gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa
     3841 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa
     3901 tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc gggcaggacg
     3961 cccgccataa actgccaggc atcaaattaa gcagaaggcc atcctgacgg atggcctttt
     4021 tgcgtttcta caaactcctc gagttcaaat atgtatccgc tcatgagaca ataaccctga
     4081 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     4141 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     4201 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     4261 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     4321 tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca agagcaactc
     4381 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     4441 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     4501 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     4561 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     4621 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     4681 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     4741 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     4801 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     4861 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     4921 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     4981 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5041 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5101 ttccactgag cga
//
