LOCUS       sequence-276081-        7670 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for phycocyanobilin (PCB) synthesis in mammalian 
            cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7670)
  AUTHORS   Uda Y, Miura H, Goto Y, Yamamoto K, Mii Y, Kondo Y, Takada S, Aoki K
  TITLE     Improvement of Phycocyanobilin Synthesis for Genetically Encoded 
            Phytochrome-Based Optogenetics.
  JOURNAL   ACS Chem Biol. 2020 Nov 20;15(11):2896-2906. doi: 
            10.1021/acschembio.0c00477. Epub 2020 Nov 9.
  PUBMED    33164485
REFERENCE   2  (bases 1 to 7670)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acschembio.0c00477"; journalName: "ACS Chem Biol"; date: 
            "2020-11-20- 20"; volume: "15"; issue: "11"; pages: "2896-2906"
COMMENT     Sequence Label: pCAGGS-synPCB2.2
FEATURES             Location/Qualifiers
     source          1..7670
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        389..665
                     /label=chicken beta-actin promoter
     intron          666..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1606..1628
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1740..1814
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             3045..3101
                     /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             3102..3176
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      4367..4376
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4656..4679
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4692..4711)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4757..4812
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4811..4830)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5173..5189)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5173..5189)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5186..5208)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5197..5213
                     /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(join(5221..5227,5228..5245,5246..5251))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5266..5287
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5346..5541
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5392..5527
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5547..5681
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5584..5603)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5638..5657
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5749..5766)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5920..6508)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6000..6019)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6679..7470,7471..7539))
                     /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     7302..7321
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7540..7644)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgatcgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc
      421 cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt
      481 gcagcgatgg gggcgggggg gggggggggg cgcgcncagg cggggcgggg cggggcgagg
      541 ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc
      721 gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt
      781 ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc
      841 gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggggtg
      901 cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg
      961 agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg
     1021 ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg
     1081 gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg caaccccccc
     1141 tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg
     1201 ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg
     1261 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
     1321 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
     1381 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
     1441 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
     1501 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
     1561 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1621 gtgtgaccgg cggctctagc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caagaattca
     1741 tgtccgtcct gacgccgctg ctgctgcggg gcttgacagg ctcggcccgg cggctcccag
     1801 tgccgcgcgc caagatccat tcgttgctcg agaatattta ccgacctaac aatccttgca
     1861 ttgggaaagt cattagcaat gaggagctgg tgcgagaggg gggagaaggg actgtaaaac
     1921 atataatctt cgatatatca ggcacggagc ttcggtatct tgaggggcaa tctattggaa
     1981 tcataccggc gggtactgat gccaatggaa aaccgcataa attgcgcctc tactctatag
     2041 cttctactag gcacggggat ttccaggatg ataagaccgt ctctttgtgt gtccgcaggc
     2101 tggaatacaa ggataaggaa accggagaga caatatatgg ggtatgttcc tcatacctga
     2161 atcaattgca accaggagat gaagtgaaga taacggggcc tgttggtaag gagatgctcc
     2221 tgtcagatga cccagaagct acaattataa tgctggcgac tgggactggc atagcgcctt
     2281 tccgagcctt cctctggaga atgttcaagg aaaacaaccc cgattatcaa ttcaaggggc
     2341 ttgcctggtt gttttttggt gtagcataca cggctaacat cctttacaaa gatgaactcg
     2401 aagcgattca ggcgcagtac ccggatcatt ttcgcttgac atacgccata tctagggaac
     2461 agaagacccc cgacgggggt aaaatgtata tccaagggcg aatagccgaa catgcagatg
     2521 aaatctggca actgctgcaa aagaaaaata ctcacgtcta tatgtgtggt ctcagaggaa
     2581 tggagccggg catagacgaa gcgatgaccg ctgcggcggc aaagaacggc gccgattggc
     2641 aagaatttct caagggtaca cttaagaagg aggggcgctg gcacgtcgag acgtacggcg
     2701 gatccggagg ttccggcggc tcaggaggtt ctatggctac ctataaggtc acgctggttc
     2761 gaccagatgg tagcgagact accattgatg tccctgaaga cgaatatatt ctggacgtag
     2821 ctgaggagca aggcttggat cttcctttca gctgccgggc tggtgcttgc agcacgtgcg
     2881 cgggaaaact tctggaggga gaggtcgatc agagcgatca gtccttcctc gacgatgacc
     2941 aaatagaaaa gggtttcgtt ctcacttgcg ttgcctaccc aagaagcgat tgcaaaatac
     3001 tcacaaacca ggaggaagag ctgtacggcg gccgcggctc aggtgcgacg aattttagtc
     3061 tactgaaaca agcgggagac gtggaggaaa accctggacc tatgtcagtg ctgactccac
     3121 tgctgctgag gggcctgact gggtccgcac gccgcctgcc tgtcccaaga gccaaaatcc
     3181 atagcctggg cgggtctgga ggcatgacta ccagcctggc tacaaagctg agggaaggca
     3241 ctaagaaagc tcataccatg gcagagaacg tggggtttgt ccggtgtttc ctgaagggaa
     3301 ctgtggagaa aagctcctat agaaagctgg tggcctctct gtaccacgtc tatagtgcta
     3361 tggagcagga aatggagcgc ctgaaagacc atccaatctt gggcaaaatc tactttcccg
     3421 aactgaacag gaagtctagt ctggagcgcg atctgacata ctatttcggg tccaattgga
     3481 gggaggaaat tcccccttct cccgcaactc aggcctacgt ggctcgcatc cacgaagtcg
     3541 caaacaccgc ccctgagctg ctggtggccc atagttacac acggtatctg ggagacctgt
     3601 cagggggaca gatcctgaag ggcattgctg aacgggcaat gaatctgcag gatggggagg
     3661 gaaccgcctt ctacagattc gaaagtatct cagacgagaa ggccttcaaa cagctgtatc
     3721 gacagcggct ggatgaactg cccgtggacg aggctacagc agatagaatt gtggacgagg
     3781 ctaacgctgc atttggcatg aatatgaaga ttttccagga actggagggc aacctgatcc
     3841 gggccattgg gcagctgctg tttaataccc tgacacgacg gaaacagaga ggcagcaccg
     3901 agctggccac agctgatggt ggaggtggat ctggaggtgg tggatcaatg agtctgaggc
     3961 agcaccagca tcctctgatc cagcgcctgg ccgatcgaat cgaggcaatt tggcaggcct
     4021 tctttcccct ggcaccttac gccctgccag aagacctggg ctatgtggag ggaaaactgg
     4081 agggcgaacg cctgacaatc gagaaccact gctatcaggc cccccctttt cggaagctgc
     4141 atctggaact ggctagagtg ggcgagtctc tggacattct gcactgcgtc atgtttcccg
     4201 agcctagata cgatctgccc atgttcggat gtgacctggt ggggggacgc ggacagatca
     4261 gtgccgctat tgtggatctg tcaccagtca ctggacagct gccagcagca tacacctgcg
     4321 ctctgaatgc actgcctaag ctgaccttta gacagccaag ggagctgcca ccatggggac
     4381 atatcttctc tcccttttgt atcttcatta gacctcaggg cgaagccgag gaacagcagt
     4441 tcctggatag gattggggag tatctgactc tgcactgcca gctgagccag caggcagtgc
     4501 ctaccgacca tccacaggcc gtcatcgctg gacagcggca gtattgtcag cagcagcagc
     4561 agaacgataa aactcggaga gtgctggaaa aggcttttgg cgtcccatgg gcagagagat
     4621 acatgaccac agtgctgttc gatgtgcctc cagtggacta taaagacgat gacgataaat
     4681 aagtcgacgg atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag
     4741 catctgactt ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt
     4801 ttgtgtctct cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag
     4861 tatttggttt agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg
     4921 ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga
     4981 aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta
     5041 acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta
     5101 ctcccagtca tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt
     5161 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     5221 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     5281 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg
     5341 gatccgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     5401 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     5461 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     5521 ggaggcctag gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat
     5581 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     5641 gtttgtccaa actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg
     5701 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5761 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5821 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5881 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5941 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     6001 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     6061 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6121 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6181 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6241 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6301 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6361 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6421 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6481 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6541 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6601 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6661 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6721 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6781 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6841 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6901 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6961 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     7021 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     7081 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7141 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7201 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7261 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7321 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7381 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7441 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7501 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7561 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7621 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
//
