LOCUS       sequence-155813-        4503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Contains bicistronic pTet-Scaffold(3xFLAG)- ShineDelgarno-mCherry 
            and TetR genes. Used for Western blots..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4503)
  AUTHORS   Hsiao V, de los Santos EL, Whitaker WR, Dueber JE, Murray RM
  TITLE     Design and implementation of a biomolecular concentration tracker.
  JOURNAL   ACS Synth Biol. 2015 Feb 20;4(2):150-61. doi: 10.1021/sb500024b. 
            Epub 2014 May 15.
  PUBMED    24847683
REFERENCE   2  (bases 1 to 4503)
  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/sb500024b";
            journalName: "ACS Synth Biol"; date: "2015-02-20- 20"; volume: "4"; 
            issue: "2"; pages: "150-61"
COMMENT     Sequence Label: pVH011
FEATURES             Location/Qualifiers
     source          1..4503
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             4..714
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(149..167)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     417..436
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             746..775
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             791..808
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(864..881)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1034..1080
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1122..1139)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1293..1881)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1373..1392)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2052..2843,2844..2912))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2675..2694
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2913..3017)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(3107..3150)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     CDS             complement(3290..3916)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     3821..3840
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        3932..3987
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    3968..3986
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             4145..4162
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4415..4480
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
ORIGIN
        1 ttcatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc
       61 aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
      121 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg
      181 cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag
      241 caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag
      301 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccttgcag
      361 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
      421 gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac
      481 ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac
      541 gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac
      601 gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
      661 gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa gtaataataa
      721 ctcgagtaat gagaagcttg ggcccgaaca aaaactcatc tcagaagagg atctgaatag
      781 cgccgtcgac catcatcatc atcatcattg agtttaaacg gtctccagct tggctgtttt
      841 ggcggatgag agaagatttt cagcctgata cagattaaat cagaacgcag aagcggtctg
      901 ataaaacaga atttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac
      961 tcagaagtga aacgccgtag cgccgatggt agtgtggggt ctccccatgc gagagtaggg
     1021 aactgccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     1081 ctgttgtttg tcggtgaact aatttctaga ctgcaggctt cctcgctcac tgactcgctg
     1141 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     1201 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     1261 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     1321 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     1381 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     1441 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     1501 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     1561 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     1621 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     1681 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     1741 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     1801 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     1861 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     1921 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     1981 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     2041 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     2101 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     2161 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     2221 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     2281 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     2341 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     2401 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     2461 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     2521 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     2581 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     2641 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     2701 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     2761 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     2821 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     2881 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     2941 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     3001 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt
     3061 attatcatga cattaaccta taaaaatagg cgtatcacga ggcagaattt cagataaaaa
     3121 aaatccttag ctttcgctaa ggatgatttc tggaattcac tagtgatccg tttccattta
     3181 ggtgggtacg ttggagccgc attattttcg ctttatgaat ctaaagggtg gttaactcga
     3241 catcttggtt accgtgaagt taccatcacg gaaaaaggtt atgctgcttt taagacccac
     3301 tttcacattt aagttgtttt tctaatccgc atatgatcaa ttcaaggccg aataagaagg
     3361 ctggctctgc accttggtga tcaaataatt cgatagcttg tcgtaataat ggcggcatac
     3421 tatcagtagt aggtgtttcc ctttcttctt tagcgacttg atgctcttga tcttccaata
     3481 cgcaacctaa agtaaaatgc cccacagcgc tgagtgcata taatgcattc tctagtgaaa
     3541 aaccttgttg gcataaaaag gctaattgat tttcgagagt ttcatactgt ttttctgtag
     3601 gccgtgtacc taaatgtact tttgctccat cgcgatgact tagtaaagca catctaaaac
     3661 ttttagcgtt attacgtaaa aaatcttgcc agctttcccc ttctaaaggg caaaagtgag
     3721 tatggtgcct atctaacatc tcaatggcta aggcgtcgag caaagcccgc ttatttttta
     3781 catgccaata caatgtaggc tgctctacac ctagcttctg ggcgagttta cgggttgtta
     3841 aaccttcgat tccgacctca ttaagcagct ctaatgcgct gttaatcact ttacttttat
     3901 ctaatcttga catcattaat tcctaatttt tgttgacact ctatcgttga tagagttatt
     3961 ttaccactcc ctatcagtga tagagaaaag tgaaaatcca tatgactagc agatccacta
     4021 gagtcgacta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     4081 ggccctttcg tcttcacgct agcggtacct gaagtgtggc ccaacgcaaa ctaagagatc
     4141 tatgcatcac catcaccatc acgactacga catcccgact accgaaaacc tgtacttcca
     4201 gcctccacct gcattaccgc caaagagaag gcgtggatct gggtccggca gcggtggatc
     4261 tatcacaatt cgggcagctt ttctggagaa ggaaaacacc gcactgcgta ctgagattgc
     4321 tgaactggag aaagaagtag gccgctgcga gaatattgtt agcaaatatg aaacccgcta
     4381 cggccccctg ggatccggta gtggcagcgg cagcgactac aaggaccatg acggtgatta
     4441 caaggatcat gacatcgact acaaggatga cgatgacaag tagtgaggat cctaaggagg
     4501 gaa
//
