LOCUS       sequence-220784-        7601 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NanoLuc Gateway shuttle vector for C-terminal fusions.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7601)
  AUTHORS   Trepte P, Kruse S, Kostova S, Hoffmann S, Buntru A, Tempelmeier A, 
            Secker C, Diez L, Schulz A, Klockmeier K, Zenkner M, Golusik S, Rau 
            K, Schnoegl S, Garner CC, Wanker EE
  TITLE     LuTHy: a double-readout bioluminescence-based two-hybrid technology 
            for quantitative mapping of protein-protein interactions in 
            mammalian cells.
  JOURNAL   Mol Syst Biol. 2018 Jul 11;14(7):e8071. doi: 10.15252/msb.20178071.
  PUBMED    29997244
REFERENCE   2  (bases 1 to 7601)
  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: "Mol Syst 
            Biol."; date: "2018-07-11"; volume: "14(7):e8071. doi"; pages: " 
            10.15252/msb.20178071"
COMMENT     Sequence Label: pcDNA3.1 GW-NL-myc
FEATURES             Location/Qualifiers
     source          1..7601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             142..447
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     326..345
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(488..612)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             620..1129
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     CDS             1130..1159
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1200..1217)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1206..1430
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1476..1904
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1563..1582)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1773..1794
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1913..1933)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1918..2247
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2098..2233
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2160..2179
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2314..3108
                     /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(2368..2387)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2978..2997
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3282..3403
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3319..3338)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3373..3392
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3452..3468)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3452..3468)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3465..3487)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3476..3492
                     /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(3500..3506,3507..3524,3525..3530))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3545..3566
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3683..3700)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3854..4442)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3934..3953)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4613..5404,5405..5473))
                     /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     5236..5255
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5474..5578)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5653..5672)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5844..6223
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6224..6427
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6378..6398
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     6472..6491
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6472..6490
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6510..6634
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(6659..6664,6665..6682,6683..6689)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             6743..7423
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     complement(6809..6828)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 tggtaagcac aaccatgcag aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg
       61 aaaatcagga agggatggct gaggtcgccc ggtttattga aatgaacggc tcttttgctg
      121 acgagaacag gggctggtga aatgcagttt aaggtttaca cctataaaag agagagccgt
      181 tatcgtctgt ttgtggatgt acagagtgat attattgaca cgcccgggcg acggatggtg
      241 atccccctgg ccagtgcacg tctgctgtca gataaagtcc cccgtgaact ttacccggtg
      301 gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag tgtgccggtc
      361 tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat caaaaacgcc
      421 attaacctga tgttctgggg aatataaatg tcaggctccc ttatacacag ccagtctgca
      481 ggtcgaccat agtgactgga tatgttgtgt tttacagtat tatgtagtct gttttttatg
      541 caaaatctaa tttaatatat tgatatttat atcattttac gtttctcgtt cagctttctt
      601 gtacaaagtg gtgaagcttg tcttcacact cgaagatttc gttggggact ggcgacagac
      661 agccggctac aacctggacc aagtccttga acagggaggt gtgtccagtt tgtttcagaa
      721 tctcggggtg tccgtaactc cgatccaaag gattgtcctg agcggtgaaa atgggctgaa
      781 gatcgacatc catgtcatca tcccgtatga aggtctgagc ggcgaccaaa tgggccagat
      841 cgaaaaaatt tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgca
      901 ctatggcaca ctggtaatcg acggggttac gccgaacatg atcgactatt tcggacggcc
      961 gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa
     1021 cggcaacaaa attatcgacg agcgcctgat caaccccgac ggctccctgc tgttccgagt
     1081 aaccatcaac ggagtgaccg gctggcggct gtgcgaacgc attctggcgg aacagaaact
     1141 gatctctgaa gaagacctgt gactcgagtc tagagggccc gtttaaaccc gctgatcagc
     1201 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     1261 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     1321 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     1381 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     1441 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     1501 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1561 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1621 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     1681 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     1741 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     1801 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     1861 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     1921 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     1981 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     2041 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     2101 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     2161 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     2221 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     2281 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     2341 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     2401 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     2461 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     2521 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     2581 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     2641 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     2701 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     2761 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     2821 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     2881 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     2941 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3001 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3061 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3121 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3181 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3241 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     3301 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3361 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     3421 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     3481 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3541 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3601 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3661 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3721 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3781 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3841 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3901 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3961 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4021 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4081 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4141 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4201 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4261 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4321 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4381 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4441 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4501 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4561 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4621 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4681 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4741 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4801 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4861 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4921 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4981 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5041 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5101 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5161 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5221 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5281 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5341 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5401 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5461 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5521 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5581 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     5641 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     5701 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     5761 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     5821 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
     5881 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     5941 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     6001 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     6061 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     6121 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     6181 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     6241 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     6301 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     6361 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     6421 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
     6481 tcactatagg gagacccaag ctggctagca caagtttgta caaaaaagct gaacgagaaa
     6541 cgtaaaatga tataaatatc aatatattaa attagatttt gcataaaaaa cagactacat
     6601 aatactgtaa aacacaacat atccagtcac tatggcggcc gcattaggca ccccaggctt
     6661 tacactttat gcttccggct cgtataatgt gtggattttg agttaggatc cgtcgagatt
     6721 ttcaggagct aaggaagcta aaatggagaa aaaaatcact ggatatacca ccgttgatat
     6781 atcccaatgg catcgtaaag aacattttga ggcatttcag tcagttgctc aatgtaccta
     6841 taaccagacc gttcagctgg atattacggc ctttttaaag accgtaaaga aaaataagca
     6901 caagttttat ccggccttta ttcacattct tgcccgcctg atgaatgctc atccggaatt
     6961 ccgtatggca atgaaagacg gtgagctggt gatatgggat agtgttcacc cttgttacac
     7021 cgttttccat gagcaaactg aaacgttttc atcgctctgg agtgaatacc acgacgattt
     7081 ccggcagttt ctacacatat attcgcaaga tgtggcgtgt tacggtgaaa acctggccta
     7141 tttccctaaa gggtttattg agaatatgtt tttcgtctca gccaatccct gggtgagttt
     7201 caccagtttt gatttaaacg tggccaatat ggacaacttc ttcgcccccg ttttcaccat
     7261 gggcaaatat tatacgcaag gcgacaaggt gctgatgccg ctggcgattc aggttcatca
     7321 tgccgtttgt gatggcttcc atgtcggcag aatgcttaat gaattacaac agtactgcga
     7381 tgagtggcag gcggggcgta atctagagga tccggcttac taaaagccag ataacagtat
     7441 gcgtatttgc gcgctgattt ttgcggtata agaatatata ctgatatgta tacccgaagt
     7501 atgtcaaaaa gaggtatgct atgaagcagc gtattacagt gacagttgac agcgacagct
     7561 atcagttgct caaggcatat atgatgtcaa tatctccggt c
//
