LOCUS       sequence-193717-        5767 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses PIF3 (1-100 aa) fused with mEGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5767)
  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 5767)
  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"; journalName: "Proc Natl 
            Acad Sci U S A. 2017 Oct 24. pii: 201707190. doi: 
            10.1073/pnas.1707190114."
COMMENT     Sequence Label: pCAGGS-PIF3-mEGFP
FEATURES             Location/Qualifiers
     source          1..5767
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     intron          147..1167
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1087..1109
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1175..1194
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             join(1558..1560,1561..1563,1564..2277)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELSR"
     primer_bind     complement(1603..1624)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1864..1883)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2211..2232
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2287..2306)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2352..2407
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2406..2425)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2768..2784)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2768..2784)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2781..2803)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2792..2808
                     /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(2816..2822,2823..2840,2841..2846))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2861..2882
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2941..3136
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2987..3122
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3142..3276
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3179..3198)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3233..3252
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3344..3361)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3515..4103)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3595..3614)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4274..5065,5066..5134))
                     /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     4897..4916
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5135..5239)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5274..5653
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggggggcggg gcggggcgag gggcggggcg gggcgaggcg gagaggtgcg gcggcagcca
       61 atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct
      121 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc
      181 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg
      241 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc
      301 ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg
      361 gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct
      421 ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca
      481 gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg
      541 ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt
      601 cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac ggcccggctt
      661 cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc
      721 ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggagggc tcgggggagg
      781 ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc agccattgcc
      841 ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc tgtgcggagc
      901 cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg
      961 ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc
     1021 tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca
     1081 gggcggggtt cggcttctgg cgtgtgaccg gcggctctag ctagagcctc tgctaaccat
     1141 gttcatgcct tcttcttttt cctacagctc ctgggcaacg tgctggttat tgtgctgtct
     1201 catcattttg gcaaagaatt caccatgcct ctgtttgagc ttttcaggct caccaaagct
     1261 aagcttgaat ctgctcaaga caggaaccct tctccacctg tagatgaagt tgtggagctg
     1321 gtgtgggaaa atggtcagat atcaactcaa agtcagtcaa gtagatcgag gaacattcct
     1381 ccaccacaag caaactcttc tagagctaga gagattggaa atggctcaaa gacgactatg
     1441 gtggacgaga tccctatgtc agtgccatca ctaatgacgg gtttgagtca agacgatgac
     1501 tttgttccat ggttgaatca tcatctcgag ggaggatccg gaggttccgg cggccgcatg
     1561 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1621 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1681 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1741 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1801 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1861 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1921 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1981 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     2041 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     2101 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     2161 ctgagcaccc agtccaagct tagcaaagac cccaacgaga agcgcgatca catggtcctg
     2221 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgtc tagataagtc
     2281 gacggatctt tttccctctg ccaaaaatta tggggacatc atgaagcccc ttgagcatct
     2341 gacttctggc taataaagga aatttatttt cattgcaata gtgtgttgga attttttgtg
     2401 tctctcactc ggaaggacat atgggagggc aaatcattta aaacatcaga atgagtattt
     2461 ggtttagagt ttggcaacat atgcccatat gctggctgcc atgaacaaag gttggctata
     2521 aagaggtcat cagtatatga aacagccccc tgctgtccat tccttattcc atagaaaagc
     2581 cttgacttga ggttagattt tttttatatt ttgttttgtg ttattttttt ctttaacatc
     2641 cctaaaattt tccttacatg ttttactagc cagatttttc ctcctctcct gactactccc
     2701 agtcatagct gtccctcttc tcttatggag atccctcgac ctgcagccca agcttggcgt
     2761 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     2821 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     2881 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcc
     2941 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     3001 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     3061 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     3121 cctaggcttt tgcaaaaagc taacttgttt attgcagctt ataatggtta caaataaagc
     3181 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3241 tccaaactca tcaatgtatc ttatcatgtc tggatccgct gcattaatga atcggccaac
     3301 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     3361 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3421 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     3481 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3541 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3601 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3661 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3721 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     3781 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     3841 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     3901 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     3961 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4021 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4081 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4141 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     4201 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     4261 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4321 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4381 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     4441 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     4501 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     4561 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     4621 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     4681 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     4741 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     4801 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     4861 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     4921 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     4981 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     5041 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     5101 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     5161 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     5221 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgggtcg atcgacattg
     5281 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     5341 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     5401 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     5461 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     5521 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     5581 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     5641 cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc
     5701 ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag cgatgggggc
     5761 ggggggg
//
