LOCUS       62056_2590        9817 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9817)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..9817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    37..840
                     /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    847..872
                     /label=SA
                     /note="splice acceptor site"
     CDS             896..949
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             959..1555
                     /codon_start=1
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    1596..1820
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     intron          2484..3492
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and
                     rabbit beta-globin"
     CDS             3557..4273
                     /codon_start=1
                     /label=hrGFP
                     /note="humanized Renilla green fluorescent protein"
                     /translation="MVSKQILKNTGLQEIMSFKVNLEGVVNNHVFTMEGCGKGNILFGN
                     QLVQIRVTKGAPLPFAFDILSPAFQYGNRTFTKYPEDISDFFIQSFPAGFVYERTLRYE
                     DGGLVEIRSDINLIEGMFVYRVEYKGRNFPNDGPVMKKTITGLQPSFEVVYMNDGVLVG
                     QVILVYRLNSGKFYSCHMRTLMKSKGVVKDFPEYHFIQHRLEKTYVEDGGFVEQHETAI
                     AQLTSLGKPLGSLHEWV"
     polyA_signal    4443..4498
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4859..4875)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4883..4899)
                     /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(4907..4937)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4952..4973)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    5047..5883
                     /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(5924..5942)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5949..5965)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             6103..6402
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     CDS             complement(7805..8662)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      8786..9374
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    9662..9683
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        9698..9728
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9736..9752
                     /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)."
     primer_bind     9760..9776
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        9797..9815
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 ctagctagtc ctgcaggttt aaacgaattc gccctttgct ttctctgacc agcattctct
       61 cccctgggcc tgtgccgctt tctgtctgca gcttgtggcc tgggtcacct ctacggctgg
      121 cccagatcct tccctgccgc ctccttcagg ttccgtcttc ctccactccc tcttcccctt
      181 gctctctgct gtgttgctgc ccaaggatgc tctttccgga gcacttcctt ctcggcgctg
      241 caccacgtga tgtcctctga gcggatcctc cccgtgtctg ggtcctctcc gggcatctct
      301 cctccctcac ccaaccccat gccgtcttca ctcgctgggt tcccttttcc ttctccttct
      361 ggggcctgtg ccatctctcg tttcttagga tggccttctc cgacggatgt ctcccttgcg
      421 tcccgcctcc ccttcttgta ggcctgcatc atcaccgttt ttctggacaa ccccaaagta
      481 ccccgtctcc ctggctttag ccacctctcc atcctcttgc tttctttgcc tggacacccc
      541 gttctcctgt ggattcgggt cacctctcac tcctttcatt tgggcagctc ccctaccccc
      601 cttacctctc tagtctgtgc tagctcttcc agccccctgt catggcatct tccaggggtc
      661 cgagagctca gctagtcttc ttcctccaac ccgggcccct atgtccactt caggacagca
      721 tgtttgctgc ctccagggat cctgtgtccc cgagctggga ccaccttata ttcccagggc
      781 cggttaatgt ggctctggtt ctgggtactt ttatctgtcc cctccacccc acagtggggc
      841 aagcttctga cctcttctct tcctcccaca gggcctcgag agatctggca gcggagaggg
      901 cagaggaagt cttctaacat gcggtgacgt ggaggagaat cccggcccta ggctcgagat
      961 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg
     1021 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg
     1081 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     1141 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     1201 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     1261 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     1321 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     1381 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     1441 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     1501 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgatc
     1561 tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     1621 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1681 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     1741 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     1801 ggatgcggtg ggctctatgg gtctcgacat tgattattga ctagttatta atagtaatca
     1861 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     1921 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     1981 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga ctatttacgg
     2041 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     2101 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     2161 cctacttggc agtacatcta cgtattagtc atcgctatta ccatgggtcg aggtgagccc
     2221 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
     2281 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggcgc gcgccaggcg
     2341 gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg gcagccaatc
     2401 agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata
     2461 aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgttgcc ttcgccccgt gccccgctcc
     2521 gcgccgcctc gcgccgcccg ccccggctct gactgaccgc gttactccca caggtgagcg
     2581 ggcgggacgg cccttctcct ccgggctgta attagcgctt ggtttaatga cggctcgttt
     2641 cttttctgtg gctgcgtgaa agccttaaag ggctccggga gggccctttg tgcggggggg
     2701 agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga gcgccgcgtg cggcccgcgc
     2761 tgcccggcgg ctgtgagcgc tgcgggcgcg gcgcggggct ttgtgcgctc cgcgtgtgcg
     2821 cgaggggagc gcggccgggg gcggtgcccc gcggtgcggg ggggctgcga ggggaacaaa
     2881 ggctgcgtgc ggggtgtgtg cgtggggggg tgagcagggg gtgtgggcgc ggcggtcggg
     2941 ctgtaacccc cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc
     3001 ggggctccgt gcggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt
     3061 gggggtgccg ggcggggcgg ggccgcctcg ggccggggag ggctcggggg aggggcgcgg
     3121 cggccccgga gcgccggcgg ctgtcgaggc gcggcgagcc gcagccattg ccttttatgg
     3181 taatcgtgcg agagggcgca gggacttcct ttgtcccaaa tctggcggag ccgaaatctg
     3241 ggaggcgccg ccgcaccccc tctagcgggc gcgggcgaag cggtgcggcg ccggcaggaa
     3301 ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc tccatctcca
     3361 gcctcggggc tgccgcaggg ggacggctgc cttcgggggg gacggggcag ggcggggttc
     3421 ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc
     3481 tttttcctac agctcctggg caacgtgctg gttgttgtgc tgtctcatca ttttggcaaa
     3541 gaattcgtcg accaccatgg tgagcaagca gatcctgaag aacaccggcc tgcaggagat
     3601 catgagcttc aaggtgaacc tggagggcgt ggtgaacaac cacgtgttca ccatggaggg
     3661 ctgcggcaag ggcaacatcc tgttcggcaa ccagctggtg cagatccgcg tgaccaaggg
     3721 cgcccccctg cccttcgcct tcgacatcct gagccccgcc ttccagtacg gcaaccgcac
     3781 cttcaccaag taccccgagg acatcagcga cttcttcatc cagagcttcc ccgccggctt
     3841 cgtgtacgag cgcaccctgc gctacgagga cggcggcctg gtggagatcc gcagcgacat
     3901 caacctgatc gaggggatgt tcgtgtaccg cgtggagtac aagggccgca acttccccaa
     3961 cgacggcccc gtgatgaaga agaccatcac cggcctgcag cccagcttcg aggtggtgta
     4021 catgaacgac ggcgtgctgg tgggccaggt gatcctggtg taccgcctga acagcggcaa
     4081 gttctacagc tgccacatgc gcaccctgat gaagagcaag ggcgtggtga aggacttccc
     4141 cgagtaccac ttcatccagc accgcctgga gaagacctac gtggaggacg gcggcttcgt
     4201 agagcagcac gagaccgcca tcgcccagct gaccagcctg ggcaagcccc tgggcagcct
     4261 gcacgagtgg gtgtaaatcg atgatatcac gcgtgaattc actcctcagg tgcaggctgc
     4321 ctatcagaag gtggtggctg gtgtggccaa tgccctggct cacaaatacc actgagatct
     4381 ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg
     4441 ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact
     4501 cggaaggaca tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag
     4561 tttggcaaca tatgcccata tgctggctgc catgaacaaa ggttggctat aaagaggtca
     4621 tcagtatatg aaacagcccc ctgctgtcca ttccttattc catagaaaag ccttgacttg
     4681 aggttagatt ttttttatat tttgttttgt gttatttttt tctttaacat ccctaaaatt
     4741 ttccttacat gttttactag ccagattttt cctcctctcc tgactactcc cagtcatagc
     4801 tgtccctctt ctcttatgga gatccctcga cctgcagccc aagcttggcg taatcatggt
     4861 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4921 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4981 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagcggatc gacagtacta
     5041 agctttacta gggacaggat tggtgacaga aaagccccat ccttaggcct cctccttcct
     5101 agtctcctga tattgggtct aacccccacc tcctgttagg cagattcctt atctggtgac
     5161 acacccccat ttcctggagc catctctctc cttgccagaa cctctaaggt ttgcttacga
     5221 tggagccaga gaggatcctg ggagggagag cttggcaggg ggtgggaggg aaggggggga
     5281 tgcgtgacct gcccggttct cagtggccac cctgcgctac cctctcccag aacctgagct
     5341 gctctgacgc ggctgtctgg tgcgtttcac tgatcctggt gctgcagctt ccttacactt
     5401 cccaagagga gaagcagttt ggaaaaacaa aatcagaata agttggtcct gagttctaac
     5461 tttggctctt cacctttcta gtccccaatt tatattgttc ctccgtgcgt cagttttacc
     5521 tgtgagataa ggccagtagc cagccccgtc ctggcagggc tgtggtgagg aggggggtgt
     5581 ccgtgtggaa aactcccttt gtgagaatgg tgcgtcctag gtgttcacca ggtcgtggcc
     5641 gcctctactc cctttctctt tctccatcct tctttcctta aagagtcccc agtgctatct
     5701 gggacatatt cctccgccca gagcagggtc ccgcttccct aaggccctgc tctgggcttc
     5761 tgggtttgag tccttggcaa gcccaggaga ggcgctcagg cttccctgtc ccccttcctc
     5821 gtccaccatc tcatgcccct ggctctcctg ccccttccct acaggggttc ctggctctgc
     5881 tctaagggca agggcgaatt cgcggccgct aaattcaatt cgccctatag tgagtcgtat
     5941 tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     6001 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     6061 accgatcgcc cttcccaaca gttgcgcagc ctatacgtac ggcagtttaa ggtttacacc
     6121 tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg
     6181 ccggggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc
     6241 cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat
     6301 atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa
     6361 aatgacatca aaaacgccat taacctgatg ttctggggaa tataaatgtc aggcatgaga
     6421 ttatcaaaaa ggatcttcac ctagatcctt ttcacgtaga aagccagtcc gcagaaacgg
     6481 tgctgacccc ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca
     6541 aagagaaagc aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta
     6601 tggacagcaa gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc
     6661 tgcaaagtaa actggatggc tttcttgccg ccaaggatct gatggcgcag gggatcaagc
     6721 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     6781 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     6841 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     6901 aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg
     6961 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     7021 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     7081 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     7141 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     7201 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     7261 ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatgca
     7321 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     7381 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     7441 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     7501 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaattattaa
     7561 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     7621 catcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     7681 tacattcaaa tatgtatccg ctcatgagat tatcaaaaag gatcttcacc tagatccttt
     7741 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     7801 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     7861 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     7921 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     7981 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     8041 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     8101 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     8161 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     8221 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     8281 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     8341 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     8401 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     8461 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     8521 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     8581 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     8641 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     8701 gttattgtct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     8761 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     8821 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     8881 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     8941 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     9001 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     9061 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     9121 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     9181 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     9241 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     9301 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     9361 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     9421 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     9481 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     9541 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     9601 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     9661 ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc
     9721 gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg
     9781 attacgccaa gctcagaatt aaccctcact aaaggga
//
