LOCUS       40924_42159        7288 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pT1_T1_83bp-mKate2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7288)
  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  JOURNAL   Unpublished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 7288)
  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 7288)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7288)
  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            "Unpublished"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..7288
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..95
                     /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)"
     misc_feature    129..188
                     /label=CMVmini
                     /note="CMVmini"
     protein_bind    236..256
                     /label=attB1
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     CDS             298..315
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             316..1011
                     /codon_start=1
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHK"
     protein_bind    complement(1054..1078)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1233..1288
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1649..1665)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1673..1689
                     /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(1697..1727)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1742..1763
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    1936..2772
                     /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(2802..2820)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2841..2857)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2865..2881)
                     /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(2889..2919)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(2934..2955)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3243..3831)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4005..4862)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4863..4967)
                     /label=AmpR promoter
     rep_origin      complement(4993..5448)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     5590..5606
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5616..5634
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    5649..6452
                     /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    6531..6556
                     /label=SA
                     /note="splice acceptor site"
     protein_bind    7221..7241
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
ORIGIN
        1 cggagtactg tcctccgagc ggagtactgt cctccgagcg gagtactgtc ctccgagcgg
       61 agtactgtcc tccgagcgga gttctgtcct ccgagcggag actctagata cgactcacta
      121 tagaattcta ggcgtgtacg gtgggaggcc tatataagca gagctcgttt agtgaaccgt
      181 cagatcgcct cgagtaggtt gtgtctacga ctcactatag gatccaattc gaccccaagt
      241 ttgtacaaaa aagcagggat cccggtaccc tcgacacctg ctgacggagc caccatgcac
      301 catcaccatc accatatggt gagcgagctg attaaggaga acatgcacat gaagctgtac
      361 atggagggca ccgtgaacaa ccaccacttc aagtgcacat ccgagggcga aggcaagccc
      421 tacgagggca cccagaccat gagaatcaag gcggtcgagg gcggccctct ccccttcgcc
      481 ttcgacatcc tggctaccag cttcatgtac ggcagcaaaa ccttcatcaa ccacacccag
      541 ggcatccccg acttctttaa gcagtccttc cccgagggct tcacatggga gagagtcacc
      601 acatacgaag atgggggcgt gctgaccgct acccaggaca ccagcctcca ggacggctgc
      661 ctcatctaca acgtcaagat cagaggggtg aacttcccat ccaacggccc tgtgatgcag
      721 aagaaaacac tcggctggga ggcctccacc gagacactgt accccgctga cggcggcctg
      781 gaaggcagag ccgacatggc cctgaagctc gtgggcgggg gccacctgat ctgcaacctt
      841 aagaccacat acagatccaa gaaacccgct aagaacctca agatgcccgg cgtctactat
      901 gtggacagga gactggaaag aatcaaggag gccgacaaag agacatacgt cgagcagcac
      961 gaggtggctg tggccagata ctgcgacctc cctagcaaac tggggcacaa acttaattcc
     1021 tagtaacgct cagtgcaggt gaattctacc accacccagc tttcttgtac aaagtggtac
     1081 gcgtgaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     1141 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     1201 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     1261 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     1321 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     1381 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     1441 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     1501 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     1561 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     1621 cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     1681 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     1741 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     1801 acctgtcgtg ccagcggatc gacagtacta agcttggtga attcccttat gtgagtgtaa
     1861 aaggcaggcg agtttgtccc ttaattaata gggataacag ggtaatctcg aggtttaaac
     1921 gtcgacgcgg ccgctactag ggacaggatt ggtgacagaa aagccccatc cttaggcctc
     1981 ctccttccta gtctcctgat attgggtcta acccccacct cctgttaggc agattcctta
     2041 tctggtgaca cacccccatt tcctggagcc atctctctcc ttgccagaac ctctaaggtt
     2101 tgcttacgat ggagccagag aggatcctgg gagggagagc ttggcagggg gtgggaggga
     2161 agggggggat gcgtgacctg cccggttctc agtggccacc ctgcgctacc ctctcccaga
     2221 acctgagctg ctctgacgcg gctgtctggt gcgtttcact gatcctggtg ctgcagcttc
     2281 cttacacttc ccaagaggag aagcagtttg gaaaaacaaa atcagaataa gttggtcctg
     2341 agttctaact ttggctcttc acctttctag tccccaattt atattgttcc tccgtgcgtc
     2401 agttttacct gtgagataag gccagtagcc agccccgtcc tggcagggct gtggtgagga
     2461 ggggggtgtc cgtgtggaaa actccctttg tgagaatggt gcgtcctagg tgttcaccag
     2521 gtcgtggccg cctctactcc ctttctcttt ctccatcctt ctttccttaa agagtcccca
     2581 gtgctatctg ggacatattc ctccgcccag agcagggtcc cgcttcccta aggccctgct
     2641 ctgggcttct gggtttgagt ccttggcaag cccaggagag gcgctcaggc ttccctgtcc
     2701 cccttcctcg tccaccatct catgcccctg gctctcctgc cccttcccta caggggttcc
     2761 tggctctgct ctccaccgcg gtggagctcc agcttttgtt ccctttagtg agggttaatt
     2821 gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     2881 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     2941 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     3001 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     3061 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3121 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     3181 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     3241 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     3301 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     3361 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     3421 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     3481 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     3541 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     3601 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     3661 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     3721 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     3781 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     3841 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     3901 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     3961 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4021 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4081 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4141 cgagagccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     4201 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     4261 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     4321 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     4381 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     4441 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     4501 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     4561 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     4621 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     4681 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     4741 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     4801 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     4861 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     4921 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     4981 aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt
     5041 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     5101 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     5161 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     5221 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     5281 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     5341 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     5401 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat
     5461 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     5521 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     5581 cacgacgttg taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat
     5641 tgggtacctg ctttctctga ccagcattct ctcccctggg cctgtgccgc tttctgtctg
     5701 cagcttgtgg cctgggtcac ctctacggct ggcccagatc cttccctgcc gcctccttca
     5761 ggttccgtct tcctccactc cctcttcccc ttgctctctg ctgtgttgct gcccaaggat
     5821 gctctttccg gagcacttcc ttctcggcgc tgcaccacgt gatgtcctct gagcggatcc
     5881 tccccgtgtc tgggtcctct ccgggcatct ctcctccctc acccaacccc atgccgtctt
     5941 cactcgctgg gttccctttt ccttctcctt ctggggcctg tgccatctct cgtttcttag
     6001 gatggccttc tccgacggat gtctcccttg cgtcccgcct ccccttcttg taggcctgca
     6061 tcatcaccgt ttttctggac aaccccaaag taccccgtct ccctggcttt agccacctct
     6121 ccatcctctt gctttctttg cctggacacc ccgttctcct gtggattcgg gtcacctctc
     6181 actcctttca tttgggcagc tcccctaccc cccttacctc tctagtctgt gctagctctt
     6241 ccagccccct gtcatggcat cttccagggg tccgagagct cagctagtct tcttcctcca
     6301 acccgggccc ctatgtccac ttcaggacag catgtttgct gcctccaggg atcctgtgtc
     6361 cccgagctgg gaccacctta tattcccagg gccggttaat gtggctctgg ttctgggtac
     6421 ttttatctgt cccctccacc ccacagtggg gcaagcttat taccctgtta tccctaaccg
     6481 gtctaatacc caatctctcg tcttatccag atgttttata cgccaagctt ctgacctctt
     6541 ctcttcctcc cacagggcct cgagacaatt gattaacatc gatacggtac cgagttggcg
     6601 cgcctgggag ctcacgggga cagccccccc ccaaagcccc cagggatgta attacgtccc
     6661 tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg gcgctccccc
     6721 cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt ggcacgggat
     6781 cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg ggggatacgg
     6841 ggaaaaagct ttaggctgaa agagagattt agaatgacag gcgcgcctgg ccatacatcg
     6901 atacggtacc gagttggcgc gcctgggagc tcacggggac agcccccccc caaagccccc
     6961 agggatgtaa ttacgtccct cccccgctag ggggcagcag cgagccgccc ggggctccgc
     7021 tccggtccgg cgctcccccc gcatccccga gccggcagcg tgcggggaca gcccgggcac
     7081 ggggaaggtg gcacgggatc gctttcctct gaacgcttct cgctgctctt tgagcctgca
     7141 gacacctggg gggatacggg gaaaaagctt taggctgaaa gagagattta gaatgacagg
     7201 cgcgcctggt ggcgaacgat caactttgta tagaaaagtt ggctccgaat ttctcgacag
     7261 atctcatgtg attacgccaa gctacggg
//
