LOCUS       Exported                8586 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expression vector for phycocyanobilin (PCB) synthesis in mammalian 
            cells..
ACCESSION   .
VERSION     .
KEYWORDS    pCAGGS-PHFF
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8586)
  AUTHORS   Uda Y, Goto Y, Oda S, Kohchi T, Matsuda M, Aoki K
  TITLE     Efficient synthesis of phycocyanobilin in mammalian cells for 
            optogenetic control of cell signaling.
  JOURNAL   Proc Natl Acad Sci U S A. 2017 Oct 24. pii: 201707190. doi: 
            10.1073/pnas.1707190114.
  PUBMED    29078307
REFERENCE   2  (bases 1 to 8586)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8586
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1682..1701
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1732..1806
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      2253..2262
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2542..2565
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2575..2631
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2635..2709
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2740..2766
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3490..3546
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             3547..3621
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             3652..3681
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3982..4038
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4039..4113
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     primer_bind     complement(5451..5471)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5451..5469)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5490..5506)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5608..5627)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5673..5728
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5727..5746)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6089..6105)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6113..6129
                     /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(6137..6167)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6182..6203
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6262..6457
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6308..6443
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6370..6389
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    6463..6597
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6500..6519)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6554..6573
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6665..6682)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6836..7424)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6916..6935)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7595..8455)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8218..8237
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8456..8560)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg ggcgagggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattcac catgtccgtc
     1741 ctgactccac tgctgctgag gggcctgact gggtccgcac gccgcctgcc tgtcccaaga
     1801 gccaaaatcc atagcctggg cgggtctgga ggcatgagtc tgaggcagca ccagcatcct
     1861 ctgatccagc gcctggccga tcgaatcgag gcaatttggc aggccttctt tcccctggca
     1921 ccttacgccc tgccagaaga cctgggctat gtggagggaa aactggaggg cgaacgcctg
     1981 acaatcgaga accactgcta tcaggccccc ccttttcgga agctgcatct ggaactggct
     2041 agagtgggcg agtctctgga cattctgcac tgcgtcatgt ttcccgagcc tagatacgat
     2101 ctgcccatgt tcggatgtga cctggtgggg ggacgcggac agatcagtgc cgctattgtg
     2161 gatctgtcac cagtcactgg acagctgcca gcagcataca cctgcgctct gaatgcactg
     2221 cctaagctga cctttagaca gccaagggag ctgccaccat ggggacatat cttctctccc
     2281 ttttgtatct tcattagacc tcagggcgaa gccgaggaac agcagttcct ggataggatt
     2341 ggggagtatc tgactctgca ctgccagctg agccagcagg cagtgcctac cgaccatcca
     2401 caggccgtca tcgctggaca gcggcagtac tgtcagcagc agcagcagaa cgataaaact
     2461 cggagagtgc tggaaaaggc ttttggcgtc ccatgggcag agagatacat gaccacagtg
     2521 ctgttcgatg tgcctccagt ggactataag gacgatgacg ataaaggcag cggggccacc
     2581 aacttctccc tgctgaagca ggctggcgac gtggaggaaa atccaggacc cggcatgtca
     2641 gtcctgacac ctctgctgct gcgaggactg actggaagcg ccaggcgcct gcctgtgcca
     2701 cgagcaaaaa tccactcact gggcgggagc ggaggcatgt acccctatga tgtccctgac
     2761 tacgccatga ctaccagcct ggctacaaag ctgagggaag gcactaagaa agctcatacc
     2821 atggcagaga acgtggggtt tgtccggtgt ttcctgaagg gaactgtgga gaaaagctcc
     2881 tatagaaagc tggtggcctc tctgtaccac gtctatagtg ctatggagca ggaaatggag
     2941 cgcctgaaag accatccaat cgtgggcaag atctactttc ccgaactgaa caggaagtct
     3001 agtctggagc gcgatctgac atactatttc gggtccaatt ggagggagga aattccccct
     3061 tctcccgcaa ctcaggccta cgtggctcgc atccacgaag tcgcaaacac cgcccctgag
     3121 ctgctggtgg cccatagtta cacacggtat ctgggagacc tgtcaggggg acagatcctg
     3181 aagggcattg ctgaacgggc aatgaatctg caggatgggg agggaaccgc cttctacaga
     3241 ttcgaaagta tctcagacga gaaggccttc aaacagctgt atcgacagcg gctggatgaa
     3301 ctgcccgtgg acgaggctac agcagataga attgtggacg aggctaacgc tgcatttggc
     3361 atgaatatga agatcttcca ggaactggag ggcaacctga tccgggccat tgggcagctg
     3421 ctgtttaata ccctgacacg acggaaacag agaggcagca ccgagctggc cacagctgat
     3481 ggctcagggg caaccaactt cagcctgctg aagcaggccg gagacgtgga agagaatcca
     3541 ggacctatgt ccgtcctgac cccactgctg ctgaggggac tgacaggatc tgcaagaagg
     3601 ctgccagtgc ccagagccaa aattcactcc ctgggaggct ctggaggcat ggagcagaag
     3661 ctgatcagcg aggaagatct gatggcctcc tatacagtga aactgatcac tcccgacggc
     3721 gaatcaagca ttgagtgctc cgacgatacc tacatcctgg atgcagctga ggaagcaggg
     3781 ctggacctgc cttattcttg tagggcagga gcctgcagta cttgtgcagg caagattacc
     3841 gccgggagcg tggatcagag tgaccagtca tttctggacg atgaccagat cgaagctggc
     3901 tacgtgctga catgcgtcgc atatcccact tccgattgta ctattgagac ccacaaagag
     3961 gaagacctgt acggatccgg cgctacaaac ttctctctgc tgaagcaggc aggggacgtg
     4021 gaggaaaatc ctggaccaat gagtgtcctg actcccctgc tgctgagagg actgaccgga
     4081 agcgcccgcc gactgcccgt gcctagagct aagatccata gtctgggggg atcaggcggg
     4141 atgtacagcc caggatatgt ggccaccagc agccggcagt ccgatgcagg aaacaggctg
     4201 ttcgtgtacg aggtcatcgg cctgagccag tccactatga ccgatgggct ggactatccc
     4261 attcggagat ctggaagtac attcatcact gtgcctctga aacgaatgaa tcaggagatg
     4321 aggcgcatca cccggatggg aggcaaaatc gtgtcaatta agcctctgga aggcgacagc
     4381 ccactgccac acacagaggg aattgccaag ccatcacaga gcgaagggtc cggatctgag
     4441 gctgtggcaa accctgctcc agaaagcaac aagactatga caactacccc taaggagaag
     4501 aaagccgatg acatcccagt gaacatctac aggccaaaaa ccccctatat tgggaaggtg
     4561 ctggaaaatt acccactggt ccgcgaggga gctatcggaa ccgtgcagca cctgacattt
     4621 gatctgtctg ccggcgacct gcggtatctg gaagggcaga gtatcggaat cattccaccc
     4681 ggagaggatg acaagggcaa acctcacaaa ctgaggctgt acagcattgc ctccacacgc
     4741 catggggatt tcggagatga caagactgtg agcctgtgcg tcagacagct ggaataccag
     4801 aacgaagcag gagagaccgt gcagggagtc tgctccacat acctgtgcaa tatcaaagag
     4861 ggcgatgaca tcgccattac cggcccagtg gggaaggaaa tgctgctgcc tccagatgag
     4921 gacgcaaaca ttgtcatgct ggccacaggc actgggatcg ccccctttcg ggctttcctg
     4981 tggagaatgt tcaaagaaca gcacgaggac tacaagttta aaggcctggc ctggctgatc
     5041 ttcgggattc caaagtctga gaatatcctg tataaagatg acctggaaaa gatggcagcc
     5101 gagtttcccg ataacttccg cctgacatac gctattagcc gagaacagca gaatgcagag
     5161 gggggacgaa tgtatatcca gcaccgggtg gccgagaacg ctgaggaact gtggaacctg
     5221 atgcagaatc caaaaaccca tacatacatg tgcggactga agggcatgga acccgggatc
     5281 gacgaggcct ttaccgccct ggctgaacag aatggaaagg agtggacaac tttccagaga
     5341 gagatgaaga aagaacatag atggcacgtc gagacctaca tggcttcaat gacaggagga
     5401 cagcagatgg ggtagcggcc gccaccgcgg tggagctcca gcttttgttc cctttagtga
     5461 gggttaattg cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     5521 ccgctaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     5581 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     5641 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     5701 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     5761 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     5821 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     5881 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     5941 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     6001 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     6061 cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     6121 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     6181 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     6241 acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac
     6301 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     6361 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     6421 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct
     6481 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6541 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccgc
     6601 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6661 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6721 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6781 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6841 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6901 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6961 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     7021 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     7081 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     7141 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     7201 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     7261 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     7321 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     7381 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     7441 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     7501 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     7561 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     7621 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     7681 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7741 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7801 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7861 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7921 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7981 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     8041 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     8101 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     8161 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     8221 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     8281 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     8341 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     8401 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     8461 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     8521 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     8581 cacctg
//
