LOCUS       40924_8046        8764 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCAG-mKate2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8764)
  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  JOURNAL   Unpublished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 8764)
  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 8764)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8764)
  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            "Unpublished"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..8764
                     /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"
     enhancer        4313..4616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4619..4894
                     /label=chicken beta-actin promoter
     intron          4900..5908
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and
                     rabbit beta-globin"
     protein_bind    5966..5990
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             6008..6703
                     /codon_start=1
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     misc_feature    6718..6774
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(6797..6821)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    6976..7031
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(7392..7408)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7416..7432
                     /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(7440..7470)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7485..7506
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    7679..8515
                     /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(8545..8563)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8584..8600)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(8608..8624)
                     /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(8632..8662)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(8677..8698)
                     /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 cgagttaatt aatagttatt
     4261 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
     4321 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
     4381 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
     4441 actatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
     4501 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     4561 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatgggtc
     4621 gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt
     4681 ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg ggggggggcg
     4741 cgcgccaggc ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc
     4801 ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg
     4861 gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc gagtcgctgc gttgccttcg
     4921 ccccgtgccc cgctccgcgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta
     4981 ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt
     5041 taatgacggc tcgtttcttt tctgtggctg cgtgaaagcc ttaaagggct ccgggagggc
     5101 cctttgtgcg ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc
     5161 cgcgtgcggc ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt
     5221 gcgctccgcg tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg
     5281 ctgcgagggg aacaaaggct gcgtgcgggg tgtgtgcgtg ggggggtgag cagggggtgt
     5341 gggcgcggcg gtcgggctgt aacccccccc tgcacccccc tccccgagtt gctgagcacg
     5401 gcccggcttc gggtgcgggg ctccgtgcgg ggcgtggcgc ggggctcgcc gtgccgggcg
     5461 gggggtggcg gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc ggggagggct
     5521 cgggggaggg gcgcggcggc cccggagcgc cggcggctgt cgaggcgcgg cgagccgcag
     5581 ccattgcctt ttatggtaat cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg
     5641 gcggagccga aatctgggag gcgccgccgc accccctcta gcgggcgcgg gcgaagcggt
     5701 gcggcgccgg caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc
     5761 cccttctcca tctccagcct cggggctgcc gcagggggac ggctgccttc gggggggacg
     5821 gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca
     5881 tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggttg ttgtgctgtc
     5941 tcatcatttt ggcaaagaat tcgacccaag tttgtacaaa aaagcaggct gattaccggt
     6001 cgccaccatg gtgagcgagc tgattaagga gaacatgcac atgaagctgt acatggaggg
     6061 caccgtgaac aaccaccact tcaagtgcac atccgagggc gaaggcaagc cctacgaggg
     6121 cacccagacc atgagaatca aggcggtcga gggcggccct ctccccttcg ccttcgacat
     6181 cctggctacc agcttcatgt acggcagcaa aaccttcatc aaccacaccc agggcatccc
     6241 cgacttcttt aagcagtcct tccccgaggg cttcacatgg gagagagtca ccacatacga
     6301 agacgggggc gtgctgaccg ctacccagga caccagcctc caggacggct gcctcatcta
     6361 caacgtcaag atcagagggg tgaacttccc atccaacggc cctgtgatgc agaagaaaac
     6421 actcggctgg gaggcctcca ccgagaccct gtaccccgct gacggcggcc tggaaggcag
     6481 agccgacatg gccctgaagc tcgtgggcgg gggccacctg atctgcaact tgaagaccac
     6541 atacagatcc aagaaacccg ctaagaacct caagatgccc ggcgtctact atgtggacag
     6601 aagactggaa agaatcaagg aggccgacaa agagacctac gtcgagcagc acgaggtggc
     6661 tgtggccaga tactgcgacc tccctagcaa actggggcac agatgagcgg ccgcctcaga
     6721 tctcgagctc aagcttcgaa ttctgcagtc gacggtaccg cgggcccggg atccaccgga
     6781 tctagataac tgatctaccc agctttcttg tacaaagtgg tacgcgtgaa ttcactcctc
     6841 aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg gctcacaaat
     6901 accactgaga tctttttccc tctgccaaaa attatgggga catcatgaag ccccttgagc
     6961 atctgacttc tggctaataa aggaaattta ttttcattgc aatagtgtgt tggaattttt
     7021 tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat cagaatgagt
     7081 atttggttta gagtttggca acatatgccc atatgctggc tgccatgaac aaaggttggc
     7141 tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta ttccatagaa
     7201 aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt ttttctttaa
     7261 catccctaaa attttcctta catgttttac tagccagatt tttcctcctc tcctgactac
     7321 tcccagtcat agctgtccct cttctcttat ggagatccct cgacctgcag cccaagcttg
     7381 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     7441 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     7501 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagcgg
     7561 atcgacagta ctaagcttgg tctaataccc aatctctcgt cttatccaga tgttttatac
     7621 gccttaatta atagggataa cagggtaatc tcgaggttta aacgtcgacg cggccgctac
     7681 tagggacagg attggtgaca gaaaagcccc atccttaggc ctcctccttc ctagtctcct
     7741 gatattgggt ctaaccccca cctcctgtta ggcagattcc ttatctggtg acacaccccc
     7801 atttcctgga gccatctctc tccttgccag aacctctaag gtttgcttac gatggagcca
     7861 gagaggatcc tgggagggag agcttggcag ggggtgggag ggaagggggg gatgcgtgac
     7921 ctgcccggtt ctcagtggcc accctgcgct accctctccc agaacctgag ctgctctgac
     7981 gcggctgtct ggtgcgtttc actgatcctg gtgctgcagc ttccttacac ttcccaagag
     8041 gagaagcagt ttggaaaaac aaaatcagaa taagttggtc ctgagttcta actttggctc
     8101 ttcacctttc tagtccccaa tttatattgt tcctccgtgc gtcagtttta cctgtgagat
     8161 aaggccagta gccagccccg tcctggcagg gctgtggtga ggaggggggt gtccgtgtgg
     8221 aaaactccct ttgtgagaat ggtgcgtcct aggtgttcac caggtcgtgg ccgcctctac
     8281 tccctttctc tttctccatc cttctttcct taaagagtcc ccagtgctat ctgggacata
     8341 ttcctccgcc cagagcaggg tcccgcttcc ctaaggccct gctctgggct tctgggtttg
     8401 agtccttggc aagcccagga gaggcgctca ggcttccctg tcccccttcc tcgtccacca
     8461 tctcatgccc ctggctctcc tgccccttcc ctacaggggt tcctggctct gctctccacc
     8521 gcggtggagc tccagctttt gttcccttta gtgagggtta attgcgcgct tggcgtaatc
     8581 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     8641 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     8701 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     8761 aatc
//
