LOCUS       62056_13370        6247 bp DNA     circular SYN 07-JUL-2019
DEFINITION  Cloning vector pKF-P14MM2AG5h, complete sequence.
ACCESSION   MK816961
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6247)
  AUTHORS   Farquhar KS, Charlebois DA, Szenk M, Cohen J, Nevozhay D, Balazsi G.
  TITLE     Role of network-mediated stochasticity in mammalian drug resistance
  JOURNAL   Nat Commun 10 (1), 2766 (2019)
  PUBMED    31235692
REFERENCE   2  (bases 1 to 6247)
  AUTHORS   Farquhar KS, Charlebois DA, Szenk M, Cohen J, Nevozhay D, Balazsi G.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-APR-2019) Laufer Center for Physical and Quantitative 
            Biology, Stony Brook University, 100 Nicolls Road, Stony Brook, NY 
            11790, USA
REFERENCE   3  (bases 1 to 6247)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun"; date: "2019"; volume: "10"; issue: "1"; pages: "2766"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-APR-2019) Laufer Center for Physical and Quantitative Biology, 
            Stony Brook University, 100 Nicolls Road, Stony Brook, NY 11790, 
            USA"
FEATURES             Location/Qualifiers
     source          1..6247
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        257..571
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     CDS             576..1319
                     /label=rtTA-Advanced
                     /note="improved tetracycline-controlled transactivator"
     misc_feature    1329..1385
                     /note="eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides; P2A; from porcine teschovirus 1 polyprotein"
     CDS             1386..2102
                     /label=EGFP
                     /note="enhanced GFP"
     polyA_signal    2205..2429
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    2713..2760
                     /label=FRT
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2768..3787
                     /label=HygR
                     /note="aminoglycoside phosphotransferase from E. coli"
     polyA_signal    3920..4053
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4090..4106)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4114..4130)
                     /label=lac operator
                     /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)."
     promoter        complement(4138..4168)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4183..4204)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(4492..5080)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5254..6111)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6112..6216)
                     /label=AmpR promoter
ORIGIN
        1 ggacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
       61 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
      121 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
      181 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgtcgaggc
      241 cctttcgtct tcactcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttac
      301 tccctatcag tgatagagaa cgatgtcgag tttactccct atcagtgata gagaacgtat
      361 gtcgagttta ctccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga
      421 tagagaacgt atgtcgagtt tatccctatc agtgatagag aacgtatgtc gagtttactc
      481 cctatcagtg atagagaacg tatgtcgagg taggcgtgta cggtgggagg cctatataag
      541 cagagctcgt ttagtgaacc gtcagatcgc ctggaatgtc tagactggac aagagcaaag
      601 tcataaacgg cgctctggaa ttactcaatg gagtcggtat cgaaggcctg acgacaagga
      661 aactcgctca aaagctggga gttgagcagc ctaccctgta ctggcacgtg aagaacaagc
      721 gggccctgct cgatgccctg ccaatcgaga tgctggacag gcatcatacc cacttctgcc
      781 ccctggaagg cgagtcatgg caagactttc tgcggaacaa cgccaagtca ttccgctgtg
      841 ctctcctctc acatcgcgac ggggctaaag tgcatctcgg cacccgccca acagagaaac
      901 agtacgaaac cctggaaaat cagctcgcgt tcctgtgtca gcaaggcttc tccctggaga
      961 acgcactgta cgctctgtcc gccgtgggcc actttacact gggctgcgta ttggaggaac
     1021 aggagcatca agtagcaaaa gaggaaagag agacacctac caccgattct atgcccccac
     1081 ttctgagaca agcaattgag ctgttcgacc ggcagggagc cgaacctgcc ttccttttcg
     1141 gcctggaact aatcatatgt ggcctggaga aacagctaaa gtgcgaaagc ggcgggccgg
     1201 ccgacgccct tgacgatttt gacttagaca tgctcccagc cgatgccctt gacgactttg
     1261 accttgatat gctgcctgct gacgctcttg acgattttga ccttgacatg ctccccgggg
     1321 gaagcggagc tactaacttc agcctgctga agcaggctgg agacgtggag gagaaccctg
     1381 gacctatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1441 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1501 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1561 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     1621 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1681 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1741 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1801 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     1861 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     1921 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     1981 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     2041 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     2101 agtaaggatc cactagtcca gtgtggtgga attctgcaga tatccagcac agtggcggcc
     2161 gctcgagtct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     2221 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     2281 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     2341 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     2401 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     2461 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     2521 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     2581 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     2641 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     2701 ttcacgtacc tagaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc
     2761 cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt
     2821 cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt
     2881 cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa
     2941 agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg aagtgcttga
     3001 cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac
     3061 gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg cggaggccat
     3121 ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat tcggaccgca
     3181 aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg atccccatgt
     3241 gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga
     3301 tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc acgcggattt
     3361 cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg actggagcga
     3421 ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt
     3481 ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc
     3541 gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt
     3601 tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc
     3661 cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga
     3721 tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc
     3781 aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     3841 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     3901 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     3961 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     4021 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     4081 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4141 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4201 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4261 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4321 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     4381 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     4441 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     4501 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4561 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4621 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4681 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4741 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4801 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4861 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4921 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4981 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     5041 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5101 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5161 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5221 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     5281 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     5341 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     5401 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     5461 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     5521 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     5581 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5641 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5701 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5761 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5821 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5881 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5941 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     6001 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     6061 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6121 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     6181 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     6241 tgacgtc
//
