LOCUS       sequence-198708-        7482 bp    DNA     circular SYN 15-APR-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 7482)
  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 7482)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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..7482
                     /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..3912
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3763..3898
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3825..3844
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3979..4773
                     /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(4033..4052)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4643..4662
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4947..5068
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4984..5003)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5038..5057
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5117..5133)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5117..5133)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5130..5152)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5141..5157
                     /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(5165..5171,5172..5189,5190..5195))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5210..5231
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5348..5365)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5519..6107)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5599..5618)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6278..7069,7070..7138))
                     /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     6901..6920
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7139..7243)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7318..7337)
                     /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 actatttacg
      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 tcggggcatc
     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 gggatttcgg cctattggtt aaaaaatgag
     3541 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     3601 gaaagtcccc aggctcccca ggcaggcaga agtatgcaaa gcatgcatct caattagtca
     3661 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3721 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3781 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3841 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3901 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
     3961 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     4021 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     4081 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     4141 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     4201 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     4261 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     4321 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     4381 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     4441 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     4501 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     4561 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     4621 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     4681 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     4741 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     4801 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     4861 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4921 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
     4981 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     5041 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
     5101 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     5161 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     5221 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     5281 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     5341 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5401 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     5461 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     5521 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     5581 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     5641 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     5701 tggcgctttc tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     5761 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5821 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     5881 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5941 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     6001 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     6061 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6121 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6181 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     6241 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     6301 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6361 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     6421 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     6481 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     6541 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     6601 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     6661 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     6721 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6781 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6841 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     6901 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     6961 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     7021 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     7081 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     7141 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     7201 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     7261 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtcga ctctcagtac
     7321 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
     7381 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
     7441 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cg
//
