LOCUS       sequence-218198-        6345 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ProteinA-NanoLuc hybrid fusion control expression vector (for BRET 
            and LuC normalization).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6345)
  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 6345)
  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 PA-NL
FEATURES             Location/Qualifiers
     source          1..6345
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        170..549
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        550..753
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     704..724
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     798..817
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        798..816
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             842..1015
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             1016..1189
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNGAQAPK"
     CDS             1217..1246
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1247..1759
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(1871..1888)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1877..2101
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2147..2575
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2234..2253)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2444..2465
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2584..2604)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2589..2918
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2769..2904
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2831..2850
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2985..3779
                     /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(3039..3058)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3649..3668
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3953..4074
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3990..4009)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4044..4063
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4123..4139)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4123..4139)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4136..4158)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4147..4163
                     /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(4171..4177,4178..4195,4196..4201))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4216..4237
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4354..4371)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4525..5113)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4605..4624)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5284..6075,6076..6144))
                     /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     5907..5926
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6145..6249)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6324..6343)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc
       61 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
      121 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      181 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      241 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      301 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      361 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      421 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      481 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      541 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      601 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      661 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      721 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
      781 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagcaccat
      841 ggtggacaac aaattcaaca aagaacaaca aaacgcgttc tatgagatct tacatttacc
      901 taacttaaac gaagaacaac gaaacgcctt catccaaagt ttaaaagatg acccaagcca
      961 aagcgctaac cttttagcag aagctaaaaa gctaaatgat gctcaggcgc cgaaagtaga
     1021 caacaaattc aacaaagaac aacaaaacgc gttctatgag atcttacatt tacctaactt
     1081 aaacgaagaa caacgaaacg ccttcatcca aagtttaaaa gatgacccaa gccaaagcgc
     1141 taacctttta gcagaagcta aaaagctaaa tggtgctcag gcgccgaaag tagacgcgaa
     1201 ctcggaattc accatggaac agaaactgat ctctgaagaa gacctggtct tcacactcga
     1261 agatttcgtt ggggactggc gacagacagc cggctacaac ctggaccaag tccttgaaca
     1321 gggaggtgtg tccagtttgt ttcagaatct cggggtgtcc gtaactccga tccaaaggat
     1381 tgtcctgagc ggtgaaaatg ggctgaagat cgacatccat gtcatcatcc cgtatgaagg
     1441 tctgagcggc gaccaaatgg gccagatcga aaaaattttt aaggtggtgt accctgtgga
     1501 tgatcatcac tttaaggtga tcctgcacta tggcacactg gtaatcgacg gggttacgcc
     1561 gaacatgatc gactatttcg gacggccgta tgaaggcatc gccgtgttcg acggcaaaaa
     1621 gatcactgta acagggaccc tgtggaacgg caacaaaatt atcgacgagc gcctgatcaa
     1681 ccccgacggc tccctgctgt tccgagtaac catcaacgga gtgaccggct ggcggctgtg
     1741 cgaacgcatt ctggcgtgaa agcttggtac cgagctcgga tccactagtc cagtgtggtg
     1801 gaattctgca gatatccagc acagtggcgg ccgctcgagt ctagagggcc cgtttaaacc
     1861 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1921 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1981 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     2041 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     2101 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     2161 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2221 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2281 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2341 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2401 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2461 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2521 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2581 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2641 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2701 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     2761 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2821 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     2881 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     2941 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3001 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3061 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3121 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     3181 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3241 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3301 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     3361 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3421 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3481 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     3541 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     3601 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     3661 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     3721 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     3781 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     3841 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3901 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     3961 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4021 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4081 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4141 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4201 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4261 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4321 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4381 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4441 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4501 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4561 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4621 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4681 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4741 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4801 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4861 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4921 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4981 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5041 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5101 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5161 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5221 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5281 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5341 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5401 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5461 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5521 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5581 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5641 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5701 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5761 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5821 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5881 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5941 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6001 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6061 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6121 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6181 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6241 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
     6301 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgca
//
