LOCUS       40924_42404        7291 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pT9_T9_72bp-mKate2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7291)
  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  JOURNAL   Unpublished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 7291)
  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 7291)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7291)
  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            "Unpublished"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..7291
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..21
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     protein_bind    69..163
                     /label=5X UAS
                     /note="five tandem copies of the 'ScaI site' 17-mer 
                     CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) 
                     that efficiently binds yeast Gal4 (Webster et al., 1988; 
                     Pfeiffer et al., 2010)"
     protein_bind    177..197
                     /label=TALER9 bs binding site
                     /bound_moiety="TALER9 bs"
     misc_feature    204..263
                     /label=CMVmini
                     /note="CMVmini"
     protein_bind    270..290
                     /label=TALER9 bs binding site
                     /bound_moiety="TALER9 bs"
     protein_bind    307..327
                     /label=attB1
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     CDS             369..386
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             387..1082
                     /codon_start=1
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHK"
     protein_bind    complement(1125..1149)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1304..1359
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1720..1736)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1744..1760
                     /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(1768..1798)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1813..1834
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    2007..2843
                     /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(2873..2891)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2912..2928)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2936..2952)
                     /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(2960..2990)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3005..3026)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3314..3902)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4076..4933)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4934..5038)
                     /label=AmpR promoter
     rep_origin      complement(5064..5519)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     5661..5677
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5687..5705
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    5720..6523
                     /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    6602..6627
                     /label=SA
                     /note="splice acceptor site"
ORIGIN
        1 caactttgta tagaaaagtt ggctccgaat ttctcgacag atctcatgtg attacgccaa
       61 gctacgggcg gagtactgtc ctccgagcgg agtactgtcc tccgagcgga gtactgtcct
      121 ccgagcggag tactgtcctc cgagcggagt tctgtcctcc gagcggagac tctagatata
      181 gaaccgatcc tcccattgaa ttctaggcgt gtacggtggg aggcctatat aagcagagct
      241 cgtttagtga accgtcagat cgcctcgagt atagaaccga tcctcccatt ggatccaatt
      301 cgaccccaag tttgtacaaa aaagcaggga tcccggtacc ctcgacacct gctgacggag
      361 ccaccatgca ccatcaccat caccatatgg tgagcgagct gattaaggag aacatgcaca
      421 tgaagctgta catggagggc accgtgaaca accaccactt caagtgcaca tccgagggcg
      481 aaggcaagcc ctacgagggc acccagacca tgagaatcaa ggcggtcgag ggcggccctc
      541 tccccttcgc cttcgacatc ctggctacca gcttcatgta cggcagcaaa accttcatca
      601 accacaccca gggcatcccc gacttcttta agcagtcctt ccccgagggc ttcacatggg
      661 agagagtcac cacatacgaa gatgggggcg tgctgaccgc tacccaggac accagcctcc
      721 aggacggctg cctcatctac aacgtcaaga tcagaggggt gaacttccca tccaacggcc
      781 ctgtgatgca gaagaaaaca ctcggctggg aggcctccac cgagacactg taccccgctg
      841 acggcggcct ggaaggcaga gccgacatgg ccctgaagct cgtgggcggg ggccacctga
      901 tctgcaacct taagaccaca tacagatcca agaaacccgc taagaacctc aagatgcccg
      961 gcgtctacta tgtggacagg agactggaaa gaatcaagga ggccgacaaa gagacatacg
     1021 tcgagcagca cgaggtggct gtggccagat actgcgacct ccctagcaaa ctggggcaca
     1081 aacttaattc ctagtaacgc tcagtgcagg tgaattctac caccacccag ctttcttgta
     1141 caaagtggta cgcgtgaatt cactcctcag gtgcaggctg cctatcagaa ggtggtggct
     1201 ggtgtggcca atgccctggc tcacaaatac cactgagatc tttttccctc tgccaaaaat
     1261 tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt
     1321 ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg
     1381 gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat
     1441 atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc
     1501 cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata
     1561 ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta
     1621 gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg
     1681 agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     1741 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     1801 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     1861 ccagtcggga aacctgtcgt gccagcggat cgacagtact aagcttggtc taatacccaa
     1921 tctctcgtct tatccagatg ttttatacgc cttaattaat agggataaca gggtaatctc
     1981 gaggtttaaa cgtcgacgcg gccgctacta gggacaggat tggtgacaga aaagccccat
     2041 ccttaggcct cctccttcct agtctcctga tattgggtct aacccccacc tcctgttagg
     2101 cagattcctt atctggtgac acacccccat ttcctggagc catctctctc cttgccagaa
     2161 cctctaaggt ttgcttacga tggagccaga gaggatcctg ggagggagag cttggcaggg
     2221 ggtgggaggg aaggggggga tgcgtgacct gcccggttct cagtggccac cctgcgctac
     2281 cctctcccag aacctgagct gctctgacgc ggctgtctgg tgcgtttcac tgatcctggt
     2341 gctgcagctt ccttacactt cccaagagga gaagcagttt ggaaaaacaa aatcagaata
     2401 agttggtcct gagttctaac tttggctctt cacctttcta gtccccaatt tatattgttc
     2461 ctccgtgcgt cagttttacc tgtgagataa ggccagtagc cagccccgtc ctggcagggc
     2521 tgtggtgagg aggggggtgt ccgtgtggaa aactcccttt gtgagaatgg tgcgtcctag
     2581 gtgttcacca ggtcgtggcc gcctctactc cctttctctt tctccatcct tctttcctta
     2641 aagagtcccc agtgctatct gggacatatt cctccgccca gagcagggtc ccgcttccct
     2701 aaggccctgc tctgggcttc tgggtttgag tccttggcaa gcccaggaga ggcgctcagg
     2761 cttccctgtc ccccttcctc gtccaccatc tcatgcccct ggctctcctg ccccttccct
     2821 acaggggttc ctggctctgc tctccaccgc ggtggagctc cagcttttgt tccctttagt
     2881 gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     2941 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3001 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3061 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3121 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3181 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3241 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3301 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3361 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3421 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3481 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     3541 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     3601 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     3661 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     3721 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     3781 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3841 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3901 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3961 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4021 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4081 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4141 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4201 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4261 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4321 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4381 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4441 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4501 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4561 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     4621 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     4681 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     4741 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4801 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4861 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4921 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     4981 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5041 catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     5101 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     5161 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     5221 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     5281 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     5341 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     5401 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     5461 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     5521 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     5581 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     5641 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact
     5701 atagggcgaa ttgggtacct gctttctctg accagcattc tctcccctgg gcctgtgccg
     5761 ctttctgtct gcagcttgtg gcctgggtca cctctacggc tggcccagat ccttccctgc
     5821 cgcctccttc aggttccgtc ttcctccact ccctcttccc cttgctctct gctgtgttgc
     5881 tgcccaagga tgctctttcc ggagcacttc cttctcggcg ctgcaccacg tgatgtcctc
     5941 tgagcggatc ctccccgtgt ctgggtcctc tccgggcatc tctcctccct cacccaaccc
     6001 catgccgtct tcactcgctg ggttcccttt tccttctcct tctggggcct gtgccatctc
     6061 tcgtttctta ggatggcctt ctccgacgga tgtctccctt gcgtcccgcc tccccttctt
     6121 gtaggcctgc atcatcaccg tttttctgga caaccccaaa gtaccccgtc tccctggctt
     6181 tagccacctc tccatcctct tgctttcttt gcctggacac cccgttctcc tgtggattcg
     6241 ggtcacctct cactcctttc atttgggcag ctcccctacc ccccttacct ctctagtctg
     6301 tgctagctct tccagccccc tgtcatggca tcttccaggg gtccgagagc tcagctagtc
     6361 ttcttcctcc aacccgggcc cctatgtcca cttcaggaca gcatgtttgc tgcctccagg
     6421 gatcctgtgt ccccgagctg ggaccacctt atattcccag ggccggttaa tgtggctctg
     6481 gttctgggta cttttatctg tcccctccac cccacagtgg ggcaagctta ttaccctgtt
     6541 atccctaacc ggtgcgtttt tatgcttgta gtattgtata atgtttttaa gatccaagct
     6601 tctgacctct tctcttcctc ccacagggcc tcgagacaat tgattaacat cgatacggta
     6661 ccgagttggc gcgcctggga gctcacgggg acagcccccc cccaaagccc ccagggatgt
     6721 aattacgtcc ctcccccgct agggggcagc agcgagccgc ccggggctcc gctccggtcc
     6781 ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga cagcccgggc acggggaagg
     6841 tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg cagacacctg
     6901 gggggatacg gggaaaaagc tttaggctga aagagagatt tagaatgaca ggcgcgcctg
     6961 gccatacatc gatacggtac cgagttggcg cgcctgggag ctcacgggga cagccccccc
     7021 ccaaagcccc cagggatgta attacgtccc tcccccgcta gggggcagca gcgagccgcc
     7081 cggggctccg ctccggtccg gcgctccccc cgcatccccg agccggcagc gtgcggggac
     7141 agcccgggca cggggaaggt ggcacgggat cgctttcctc tgaacgcttc tcgctgctct
     7201 ttgagcctgc agacacctgg ggggatacgg ggaaaaagct ttaggctgaa agagagattt
     7261 agaatgacag gcgcgcctgg tggcgaacga t
//
