LOCUS       40924_44028        7502 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTRE-mKate2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7502)
  AUTHORS   Li Y, Jiang Y, Chen H, Liao W, Li Z, Weiss R, Xie Z.
  TITLE     Modular construction of mammalian gene circuits using TALE 
            transcriptional repressors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7502)
  AUTHORS   Li Y, Jiang Y, Chen H, Liao W.
  TITLE     Direct Submission
  JOURNAL   Submitted (03-SEP-2014) Bioinformatics Division/Center for Synthetic
REFERENCE   3  (bases 1 to 7502)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7502)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (03-SEP-2014) Bioinformatics Division/Center for Synthetic "
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7502
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(222..810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(984..1841)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1842..1946)
                     /label=AmpR promoter
     rep_origin      complement(1972..2427)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     2569..2585
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2595..2613
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    2628..3431
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     misc_feature    3510..3535
                     /label=SA
                     /note="splice acceptor site"
     protein_bind    4207..4227
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     promoter        4242..4556
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    4629..4653
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4671..5366
                     /codon_start=1
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     misc_feature    5381..5402
                     /label=FF5
                     /note="FF5"
     misc_feature    5403..5424
                     /label=FF5
                     /note="FF5"
     misc_feature    5425..5446
                     /label=FF5
                     /note="FF5"
     misc_feature    5447..5468
                     /label=FF5
                     /note="FF5"
     protein_bind    complement(5535..5559)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    5714..5769
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6130..6146)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6154..6170
                     /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)."
     promoter        complement(6178..6208)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6223..6244
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    6417..7253
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     promoter        complement(7283..7301)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7322..7338)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(7346..7362)
                     /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(7370..7400)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(7415..7436)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
ORIGIN
        1 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
       61 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
      121 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
      181 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
      241 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
      301 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
      361 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
      421 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
      481 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
      541 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
      601 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
      661 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
      721 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
      781 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
      841 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
      901 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
      961 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     1021 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     1081 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     1141 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     1201 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     1261 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     1321 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     1381 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     1441 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     1501 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     1561 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     1621 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     1681 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     1741 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     1801 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     1861 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     1921 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa
     1981 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     2041 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     2101 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     2161 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     2221 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     2281 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     2341 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     2401 cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg
     2461 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     2521 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
     2581 ccagtgagcg cgcgtaatac gactcactat agggcgaatt gggtacctgc tttctctgac
     2641 cagcattctc tcccctgggc ctgtgccgct ttctgtctgc agcttgtggc ctgggtcacc
     2701 tctacggctg gcccagatcc ttccctgccg cctccttcag gttccgtctt cctccactcc
     2761 ctcttcccct tgctctctgc tgtgttgctg cccaaggatg ctctttccgg agcacttcct
     2821 tctcggcgct gcaccacgtg atgtcctctg agcggatcct ccccgtgtct gggtcctctc
     2881 cgggcatctc tcctccctca cccaacccca tgccgtcttc actcgctggg ttcccttttc
     2941 cttctccttc tggggcctgt gccatctctc gtttcttagg atggccttct ccgacggatg
     3001 tctcccttgc gtcccgcctc cccttcttgt aggcctgcat catcaccgtt tttctggaca
     3061 accccaaagt accccgtctc cctggcttta gccacctctc catcctcttg ctttctttgc
     3121 ctggacaccc cgttctcctg tggattcggg tcacctctca ctcctttcat ttgggcagct
     3181 cccctacccc ccttacctct ctagtctgtg ctagctcttc cagccccctg tcatggcatc
     3241 ttccaggggt ccgagagctc agctagtctt cttcctccaa cccgggcccc tatgtccact
     3301 tcaggacagc atgtttgctg cctccaggga tcctgtgtcc ccgagctggg accaccttat
     3361 attcccaggg ccggttaatg tggctctggt tctgggtact tttatctgtc ccctccaccc
     3421 cacagtgggg caagcttatt accctgttat ccctaaccgg tgcgttttta tgcttgtagt
     3481 attgtataat gtttttaaga tccaagcttc tgacctcttc tcttcctccc acagggcctc
     3541 gagacaattg catcgatggt accagtgaaa gtctgcaatg aattcgagtt ggcgcgcctg
     3601 tcattctaaa tctctctttc agcctaaagc tttttccccg tatcccccca ggtgtctgca
     3661 ggctcaaaga gcagcgagaa gcgttcagag gaaagcgatc ccgtgccacc ttccccgtgc
     3721 ccgggctgtc cccgcacgct gccggctcgg ggatgcgggg ggagcgccgg accggagcgg
     3781 agccccgggc ggctcgctgc tgccccctag cgggggaggg acgtaattac atccctgggg
     3841 gctttgggag ggggctgtcc ccgtgagctc ccaggcgcgc ctgtcattct aaatctctct
     3901 ttcagcctaa agctttttcc ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga
     3961 gaagcgttca gaggaaagcg atcccgtgcc accttccccg tgcccgggct gtccccgcac
     4021 gctgccggct cggggatgcg gggggagcgc cggaccggag cggagccccg ggcggctcgc
     4081 tgctgccccc tagcggggga gggacgtaat tacatccctg ggggctttgg gggggggctg
     4141 tccccgtgag ctcccaggcg cgccaactcg gtaccgtatc gatgtcgacg ttaacgctag
     4201 tgatatcaac tttgtataga aaagttggct ccgaatttct cgagtttact ccctatcagt
     4261 gatagagaac gtatgtcgag tttactccct atcagtgata gagaacgatg tcgagtttac
     4321 tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta
     4381 tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttatcc ctatcagtga
     4441 tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt cgaggtaggc
     4501 gtgtacggtg ggaggcctat ataagcagag ctcgtttagt gaaccgtcag atcgcctgga
     4561 gatttcgagc tcggtacccg gggatcctct agtcagctga cgcgtgctag tggcgcgccg
     4621 aattcgaccc aagtttgtac aaaaaagcag gctgattacc ggtcgccacc atggtgagcg
     4681 agctgattaa ggagaacatg cacatgaagc tgtacatgga gggcaccgtg aacaaccacc
     4741 acttcaagtg cacatccgag ggcgaaggca agccctacga gggcacccag accatgagaa
     4801 tcaaggcggt cgagggcggc cctctcccct tcgccttcga catcctggct accagcttca
     4861 tgtacggcag caaaaccttc atcaaccaca cccagggcat ccccgacttc tttaagcagt
     4921 ccttccccga gggcttcaca tgggagagag tcaccacata cgaagacggg ggcgtgctga
     4981 ccgctaccca ggacaccagc ctccaggacg gctgcctcat ctacaacgtc aagatcagag
     5041 gggtgaactt cccatccaac ggccctgtga tgcagaagaa aacactcggc tgggaggcct
     5101 ccaccgagac cctgtacccc gctgacggcg gcctggaagg cagagccgac atggccctga
     5161 agctcgtggg cgggggccac ctgatctgca acttgaagac cacatacaga tccaagaaac
     5221 ccgctaagaa cctcaagatg cccggcgtct actatgtgga cagaagactg gaaagaatca
     5281 aggaggccga caaagagacc tacgtcgagc agcacgaggt ggctgtggcc agatactgcg
     5341 acctccctag caaactgggg cacagatgag cggccgcaaa aagcactctg atttgacaat
     5401 taaagcactc tgatttgaca attaaagcac tctgatttga caattaaagc actctgattt
     5461 gacaattaaa gcttcgaatt ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc
     5521 tagataactg atctacccag ctttcttgta caaagtggta cgcgtgaatt cactcctcag
     5581 gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac
     5641 cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat
     5701 ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg
     5761 tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat
     5821 ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta
     5881 taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa
     5941 gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca
     6001 tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc
     6061 ccagtcatag ctgtccctct tctcttatgg agatccctcg acctgcagcc caagcttggc
     6121 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     6181 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     6241 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat
     6301 cgacagtact aagcttggtc taatacccaa tctctcgtct tatccagatg ttttatacgc
     6361 cttaattaat agggataaca gggtaatctc gaggtttaaa cgtcgacgcg gccgctacta
     6421 gggacaggat tggtgacaga aaagccccat ccttaggcct cctccttcct agtctcctga
     6481 tattgggtct aacccccacc tcctgttagg cagattcctt atctggtgac acacccccat
     6541 ttcctggagc catctctctc cttgccagaa cctctaaggt ttgcttacga tggagccaga
     6601 gaggatcctg ggagggagag cttggcaggg ggtgggaggg aaggggggga tgcgtgacct
     6661 gcccggttct cagtggccac cctgcgctac cctctcccag aacctgagct gctctgacgc
     6721 ggctgtctgg tgcgtttcac tgatcctggt gctgcagctt ccttacactt cccaagagga
     6781 gaagcagttt ggaaaaacaa aatcagaata agttggtcct gagttctaac tttggctctt
     6841 cacctttcta gtccccaatt tatattgttc ctccgtgcgt cagttttacc tgtgagataa
     6901 ggccagtagc cagccccgtc ctggcagggc tgtggtgagg aggggggtgt ccgtgtggaa
     6961 aactcccttt gtgagaatgg tgcgtcctag gtgttcacca ggtcgtggcc gcctctactc
     7021 cctttctctt tctccatcct tctttcctta aagagtcccc agtgctatct gggacatatt
     7081 cctccgccca gagcagggtc ccgcttccct aaggccctgc tctgggcttc tgggtttgag
     7141 tccttggcaa gcccaggaga ggcgctcagg cttccctgtc ccccttcctc gtccaccatc
     7201 tcatgcccct ggctctcctg ccccttccct acaggggttc ctggctctgc tctccaccgc
     7261 ggtggagctc cagcttttgt tccctttagt gagggttaat tgcgcgcttg gcgtaatcat
     7321 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     7381 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     7441 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     7501 tc
//
