LOCUS       40924_42149        7370 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pT1_T1_72bp-mKate2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7370)
  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  JOURNAL   Unpublished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 7370)
  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 7370)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7370)
  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            "Unpublished"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..7370
                     /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"
     protein_bind    4275..4369
                     /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    4403..4462
                     /label=CMVmini
                     /note="CMVmini"
     protein_bind    4497..4521
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4539..5234
                     /codon_start=1
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     misc_feature    5249..5270
                     /label=FF5
                     /note="FF5"
     misc_feature    5271..5292
                     /label=FF5
                     /note="FF5"
     misc_feature    5293..5314
                     /label=FF5
                     /note="FF5"
     misc_feature    5315..5336
                     /label=FF5
                     /note="FF5"
     protein_bind    complement(5403..5427)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    5582..5637
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5998..6014)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6022..6038
                     /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(6046..6076)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6091..6112
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    6285..7121
                     /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(7151..7169)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7190..7206)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(7214..7230)
                     /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(7238..7268)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(7283..7304)
                     /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 tttaccgagc tcttattggt
     3481 tttcaaactt cattgactgt gccaagcttc 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 cgacagatct catgtgatta
     4261 cgccaagcta cgggcggagt actgtcctcc gagcggagta ctgtcctccg agcggagtac
     4321 tgtcctccga gcggagtact gtcctccgag cggagttctg tcctccgagc ggagactcta
     4381 gatacgactc actatagaat tctaggcgtg tacggtggga ggcctatata agcagagctc
     4441 gtttagtgaa ccgtcagatc gcctcgagta cgactcacta taggatccaa ttcgacccaa
     4501 gtttgtacaa aaaagcaggc tgattaccgg tcgccaccat ggtgagcgag ctgattaagg
     4561 agaacatgca catgaagctg tacatggagg gcaccgtgaa caaccaccac ttcaagtgca
     4621 catccgaggg cgaaggcaag ccctacgagg gcacccagac catgagaatc aaggcggtcg
     4681 agggcggccc tctccccttc gccttcgaca tcctggctac cagcttcatg tacggcagca
     4741 aaaccttcat caaccacacc cagggcatcc ccgacttctt taagcagtcc ttccccgagg
     4801 gcttcacatg ggagagagtc accacatacg aagacggggg cgtgctgacc gctacccagg
     4861 acaccagcct ccaggacggc tgcctcatct acaacgtcaa gatcagaggg gtgaacttcc
     4921 catccaacgg ccctgtgatg cagaagaaaa cactcggctg ggaggcctcc accgagaccc
     4981 tgtaccccgc tgacggcggc ctggaaggca gagccgacat ggccctgaag ctcgtgggcg
     5041 ggggccacct gatctgcaac ttgaagacca catacagatc caagaaaccc gctaagaacc
     5101 tcaagatgcc cggcgtctac tatgtggaca gaagactgga aagaatcaag gaggccgaca
     5161 aagagaccta cgtcgagcag cacgaggtgg ctgtggccag atactgcgac ctccctagca
     5221 aactggggca cagatgagcg gccgcaaaaa gcactctgat ttgacaatta aagcactctg
     5281 atttgacaat taaagcactc tgatttgaca attaaagcac tctgatttga caattaaagc
     5341 ttcgaattct gcagtcgacg gtaccgcggg cccgggatcc accggatcta gataactgat
     5401 ctacccagct ttcttgtaca aagtggtacg cgtgaattca ctcctcaggt gcaggctgcc
     5461 tatcagaagg tggtggctgg tgtggccaat gccctggctc acaaatacca ctgagatctt
     5521 tttccctctg ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc
     5581 taataaagga aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc
     5641 ggaaggacat atgggagggc aaatcattta aaacatcaga atgagtattt ggtttagagt
     5701 ttggcaacat atgcccatat gctggctgcc atgaacaaag gttggctata aagaggtcat
     5761 cagtatatga aacagccccc tgctgtccat tccttattcc atagaaaagc cttgacttga
     5821 ggttagattt tttttatatt ttgttttgtg ttattttttt ctttaacatc cctaaaattt
     5881 tccttacatg ttttactagc cagatttttc ctcctctcct gactactccc agtcatagct
     5941 gtccctcttc tcttatggag atccctcgac ctgcagccca agcttggcgt aatcatggtc
     6001 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     6061 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     6121 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcg acagtactaa
     6181 gcttggtgcg tttttatgct tgtagtattg tataatgttt ttaagatcct taattaatag
     6241 ggataacagg gtaatctcga ggtttaaacg tcgacgcggc cgctactagg gacaggattg
     6301 gtgacagaaa agccccatcc ttaggcctcc tccttcctag tctcctgata ttgggtctaa
     6361 cccccacctc ctgttaggca gattccttat ctggtgacac acccccattt cctggagcca
     6421 tctctctcct tgccagaacc tctaaggttt gcttacgatg gagccagaga ggatcctggg
     6481 agggagagct tggcaggggg tgggagggaa gggggggatg cgtgacctgc ccggttctca
     6541 gtggccaccc tgcgctaccc tctcccagaa cctgagctgc tctgacgcgg ctgtctggtg
     6601 cgtttcactg atcctggtgc tgcagcttcc ttacacttcc caagaggaga agcagtttgg
     6661 aaaaacaaaa tcagaataag ttggtcctga gttctaactt tggctcttca cctttctagt
     6721 ccccaattta tattgttcct ccgtgcgtca gttttacctg tgagataagg ccagtagcca
     6781 gccccgtcct ggcagggctg tggtgaggag gggggtgtcc gtgtggaaaa ctccctttgt
     6841 gagaatggtg cgtcctaggt gttcaccagg tcgtggccgc ctctactccc tttctctttc
     6901 tccatccttc tttccttaaa gagtccccag tgctatctgg gacatattcc tccgcccaga
     6961 gcagggtccc gcttccctaa ggccctgctc tgggcttctg ggtttgagtc cttggcaagc
     7021 ccaggagagg cgctcaggct tccctgtccc ccttcctcgt ccaccatctc atgcccctgg
     7081 ctctcctgcc ccttccctac aggggttcct ggctctgctc tccaccgcgg tggagctcca
     7141 gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt
     7201 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     7261 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     7321 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
//
