LOCUS       sequence-204678-        6550 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses pcDNA3 - CMVmin - 5X UAS -  pFR Luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6550)
  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 6550)
  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-118
FEATURES             Location/Qualifiers
     source          1..6550
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     621..640
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(859..963)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1038..1057)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     protein_bind    1235..1329
                     /label=5X UAS
                     /bound_moiety="GAL4"
                     /note="five tandem copies of the ""ScaI site"" 17-mer 
                     CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) 
                     that efficiently binds yeast Gal4 (Webster et al., 1988; 
                     Pfeiffer et al., 2010)"
     regulatory      1404..1413
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1410..3062
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTVLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(1476..1494)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     promoter        complement(3097..3115)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3098..3115)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3135..3152)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3141..3365
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3411..3839
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3498..3517)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3708..3729
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3848..3868)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3853..4182
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4033..4168
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4095..4114
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4249..5043
                     /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(4303..4322)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4913..4932
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5217..5338
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5254..5273)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5308..5327
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5387..5403)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5387..5403)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5400..5422)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5411..5427
                     /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(5435..5441,5442..5459,5460..5465))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5480..5501
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5618..5635)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5789..6377)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5869..5888)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6548..6550,1..789,790..858))
                     /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"
ORIGIN
        1 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
       61 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
      121 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
      181 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
      241 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
      301 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
      361 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
      421 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
      481 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
      541 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
      601 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
      661 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
      721 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
      781 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
      841 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
      901 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      961 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc
     1021 ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc
     1081 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg
     1141 caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt
     1201 cgcgaaagct tgcatgcgag tactgtcctc cgagcggagt actgtcctcc gagcggagta
     1261 ctgtcctccg agcggagtac tgtcctccga gcggagtact gtcctccgag cggagtactg
     1321 tcctccgagc ggagactcta gagggtatat aatggatccc cgggtaccga gctcgaattc
     1381 cagcttggca ttccggtact gccgccacca tggaagacgc caaaaacata aagaaaggcc
     1441 cggcgccatt ctatcctcta gaggatggaa ccgctggaga gcaactgcat aaggctatga
     1501 agagatacgc cctggttcct ggaacaattg cttttacaga tgcacatatc gaggtgaaca
     1561 tcacgtacgc ggaatacttc gaaatgtccg ttcggttggc agaagctatg aaacgatatg
     1621 ggctgaatac aaatcacaga atcgtcgtat gcagtgaaaa ctctcttcaa ttctttatgc
     1681 cggtgttggg cgcgttattt atcggagttg cagttgcgcc cgcgaacgac atttataatg
     1741 aacgtgaatt gctcaacagt atgaacattt cgcagcctac cgtagtgttt gtttccaaaa
     1801 aggggttgca aaaaattttg aacgtgcaaa aaaaattacc aataatccag aaaattatta
     1861 tcatggattc taaaacggat taccagggat ttcagtcgat gtacacgttc gtcacatctc
     1921 atctacctcc cggttttaat gaatacgatt ttgtaccaga gtcctttgat cgtgacaaaa
     1981 caattgcact gataatgaat tcctctggat ctactgtgtt acctaagggt gtggcccttc
     2041 cgcatagaac tgcctgcgtc agattctcgc atgccagaga tcctattttt ggcaatcaaa
     2101 tcattccgga tactgcgatt ttaagtgttg ttccattcca tcacggtttt ggaatgttta
     2161 ctacactcgg atatttgata tgtggatttc gagtcgtctt aatgtataga tttgaagaag
     2221 agctgttttt acgatccctt caggattaca aaattcaaag tgcgttgcta gtaccaaccc
     2281 tattttcatt cttcgccaaa agcactctga ttgacaaata cgatttatct aatttacacg
     2341 aaattgcttc tgggggcgca cctctttcga aagaagtcgg ggaagcggtt gcaaaacgct
     2401 tccatcttcc agggatacga caaggatatg ggctcactga gactacatca gctattctga
     2461 ttacacccga gggggatgat aaaccgggcg cggtcggtaa agttgttcca ttttttgaag
     2521 cgaaggttgt ggatctggat accgggaaaa cgctgggcgt taatcagaga ggcgaattat
     2581 gtgtcagagg acctatgatt atgtccggtt atgtaaacaa tccggaagcg accaacgcct
     2641 tgattgacaa ggatggatgg ctacattctg gagacatagc ttactgggac gaagacgaac
     2701 acttcttcat agttgaccgc ttgaagtctt taattaaata caaaggatat caggtggccc
     2761 ccgctgaatt ggaatcgata ttgttacaac accccaacat cttcgacgcg ggcgtggcag
     2821 gtcttcccga cgatgacgcc ggtgaacttc ccgccgccgt tgttgttttg gagcacggaa
     2881 agacgatgac ggaaaaagag atcgtggatt acgtcgccag tcaagtaaca accgcgaaaa
     2941 agttgcgcgg aggagttgtg tttgtggacg aagtaccgaa aggtcttacc ggaaaactcg
     3001 acgcaagaaa aatcagagag atcctcataa aggccaagaa gggcggaaag tccaaattgt
     3061 aagcggccgc tcgagcatgc atctagaggg ccctattcta tagtgtcacc taaatgctag
     3121 agctcgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     3181 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     3241 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     3301 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     3361 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     3421 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     3481 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     3541 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     3601 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     3661 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     3721 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     3781 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     3841 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3901 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3961 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     4021 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     4081 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     4141 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     4201 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     4261 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     4321 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4381 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     4441 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4501 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     4561 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     4621 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     4681 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     4741 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     4801 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     4861 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4921 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4981 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     5041 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     5101 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     5161 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     5221 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     5281 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     5341 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     5401 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     5461 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     5521 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5581 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     5641 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5701 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5761 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5821 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5881 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5941 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     6001 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     6061 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     6121 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     6181 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     6241 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6301 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6361 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6421 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6481 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     6541 ctgacagtta
//
