LOCUS       sequence-204872-        7481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pcDNA3 - pCMV - PIF3 - GBD, expressing PIF3 and GBD, under CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7481)
  AUTHORS   Kyriakakis P, Catanho M, Hoffner N, Thavarajah W, Hu VJ, Chao SS, 
            Hsu A, Pham V, Naghavian L, Dozier LE, Patrick GN, Coleman TP
  TITLE     Biosynthesis of Orthogonal Molecules Using Ferredoxin and 
            Ferredoxin-NADP(+) Reductase Systems Enables Genetically Encoded 
            PhyB Optogenetics.
  JOURNAL   ACS Synth Biol. 2018 Feb 16;7(2):706-717. doi: 
            10.1021/acssynbio.7b00413. Epub 2018 Jan 24.
  PUBMED    29301067
REFERENCE   2  (bases 1 to 7481)
  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/acssynbio.7b00413"; journalName: "ACS Synth Biol"; date: 
            "2018-02-16- 16"; volume: "7"; issue: "2"; pages: "706-717"
COMMENT     Sequence Label: pPKm-163
FEATURES             Location/Qualifiers
     source          1..7481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        27..406
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        407..610
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     561..581
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     655..674
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        655..673
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2292..2312
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2322..2762
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     CDS             2778..2804
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(2826..2844)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2827..2844)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2864..2881)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2870..3094
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3140..3568
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3227..3246)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3437..3458
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3577..3597)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3582..3911
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3762..3897
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3824..3843
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3978..4772
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4032..4051)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4642..4661
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4946..5067
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4983..5002)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5037..5056
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5116..5132)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5116..5132)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5129..5151)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5140..5156
                     /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(5164..5170,5171..5188,5189..5194))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5209..5230
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5347..5364)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5518..6106)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5598..5617)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6277..7068,7069..7137))
                     /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     6900..6919
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7138..7242)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7317..7336)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
       61 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      121 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      181 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      241 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      301 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      361 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
      421 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
      481 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
      541 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
      601 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
      661 gactcactat agggagaccc aagcttggta cctcgcgagc cgccaccatg cctctgtttg
      721 agcttttcag gctcaccaaa gctaagcttg aatctgctca agacaggaac ccttctccac
      781 ctgtagatga agttgtggag ctggtgtggg aaaatggtca gatatcaact caaagtcagt
      841 caagtagatc gaggaacatt cctccaccac aagcaaactc ttctagagct agagagattg
      901 gaaatggctc aaagacgact atggtggacg agatccctat gtcagtgcca tcactaatga
      961 cgggtttgag tcaagacgat gactttgttc catggttgaa tcatcatccc tcccttgatg
     1021 gatattgctc tgatttcttg cgtgatgtgt cgtctcctgt tactgtcaac gagcaagaga
     1081 gtgatatggc ggtaaaccaa actgctttcc cgttgtttca gagaagaaag gatggcaatg
     1141 aatcagctcc tgctgcttct tcgtcgcagt ataacggttt ccaatcgcat tctctgtatg
     1201 gaagtgatag agctagagat cttcctagcc aacaaaccaa tccggatcgg tttactcaga
     1261 cgcaggaacc actaattact agtaacaagc ctagtttggt caacttttca catttcttac
     1321 gccctgcaac ttttgcgaag actactaata ataaccttca tgacactaaa gaaaagagtc
     1381 ctcaaagccc gccaaatgtg tttcagacca gagttcttgg agctaaagac tctgaagata
     1441 aggttcttaa cgagtctgtt gcttctgcta cgcctaaaga taaccaaaag gcttgcctaa
     1501 tatcagagga ctcatgtaga aaagaccaag agagtgaaaa agcagttgta tgttcttctg
     1561 ttggctcggg taatagtctc gatggcccat ccgaaagtcc ttcactttct ttaaagagaa
     1621 agcattcgaa tattcaagac attgactgtc atagtgaaga tgtggaagaa gaatcaggag
     1681 atggaagaaa ggaagcaggt ccatctcgaa cgggtttggg ttcaaagaga agccgctctg
     1741 cagaagtgca taatctgtct gaaaggagac ggcgtgatag gatcaacgag aagatgcgtg
     1801 ccctgcaaga actcattcca aactgtaaca aggtggacaa agcttcgatg ctagatgaag
     1861 ccatcgagta tctcaagtca ctccaacttc aagtgcagat catgtcaatg gcgtctggtt
     1921 actatctgcc accggcggtt atgttcccac cgggtatggg gcattacccg gcagcagctg
     1981 ctgcaatggc aatgggtatg ggaatgcctt atgcaatggg cttgcctgat ttgagccgtg
     2041 gtggttcatc ggttaaccac ggaccacagt tccaagtctc ggggatgcaa caacaaccag
     2101 tggcgatggg tattccacgt gtctctggtg gtggtatctt tgccggttct tcgacgattg
     2161 gcaatggctc gactagagat ttatctggtt ctaaagatca aacaacgacg aataacaaca
     2221 gtaacttgaa accaataaag agaaaacagg ggtcttctga tcagttttgt ggatcgtcgc
     2281 cgcggccgca acctaagaaa aagagaaagg tgggcggagg catgaagcta ctgtcttcta
     2341 tcgaacaagc atgcgatatt tgccgactta aaaagctcaa gtgctccaaa gaaaaaccga
     2401 agtgcgccaa gtgtctgaag aacaactggg agtgtcgcta ctctcccaaa accaaaaggt
     2461 ctccgctgac tagggcacat ctgacagaag tggaatcaag gctagaaaga ctggaacagc
     2521 tatttctact gatttttcct cgagaagacc ttgacatgat tttgaaaatg gattctttac
     2581 aggatataaa agcattgtta acaggattat ttgtacaaga taatgtgaat aaagatgccg
     2641 tcacagatag attggcttca gtggagactg atatgcctct aacattgaga cagcatagaa
     2701 taagtgcgac atcatcatcg gaagagagta gtaacaaagg tcaaagacag ttgactgtat
     2761 cgccggaatt catggcttac ccatacgatg ttccagatta cgcttgataa tctagagggc
     2821 cctattctat agtgtcacct aaatgctaga gctcgctgat cagcctcgac tgtgccttct
     2881 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     2941 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     3001 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     3061 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     3121 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     3181 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     3241 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     3301 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     3361 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     3421 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     3481 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     3541 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     3601 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3661 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3721 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3781 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3841 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3901 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     3961 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     4021 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     4081 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     4141 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     4201 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     4261 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     4321 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     4381 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     4441 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     4501 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     4561 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4621 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4681 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4741 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     4801 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4861 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4921 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     4981 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     5041 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     5101 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     5161 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     5221 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     5281 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     5341 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5401 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5461 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5521 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5581 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     5641 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5701 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5761 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5821 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5881 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5941 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     6001 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     6061 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6121 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6181 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     6241 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6301 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6361 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6421 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6481 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6541 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6601 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6661 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6721 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6781 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6841 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6901 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6961 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     7021 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7081 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7141 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7201 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7261 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca
     7321 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
     7381 gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc
     7441 atgaagaatc tgcttagggt taggcgtttt gcgctgcttc g
//
