LOCUS       sequence-214026-        6342 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 6342)
  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 6342)
  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..6342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             907..1080
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             1081..1254
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNGAQAPK"
     CDS             1282..1311
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1312..1824
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(1936..1953)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1942..2166
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2212..2640
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2299..2318)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2509..2530
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2649..2669)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2654..2983
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2834..2969
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2896..2915
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3050..3844
                     /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(3104..3123)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3714..3733
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4018..4139
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4055..4074)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4109..4128
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4188..4204)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4188..4204)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4201..4223)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4212..4228
                     /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(4236..4242,4243..4260,4261..4266))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4281..4302
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4419..4436)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4590..5175)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4670..4689)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5346..6137,6138..6206))
                     /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     5969..5988
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6207..6311)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 accatggtgg acaacaaatt caacaaagaa caacaaaacg cgttctatga gatcttacat
      961 ttacctaact taaacgaaga acaacgaaac gccttcatcc aaagtttaaa agatgaccca
     1021 agccaaagcg ctaacctttt agcagaagct aaaaagctaa atgatgctca ggcgccgaaa
     1081 gtagacaaca aattcaacaa agaacaacaa aacgcgttct atgagatctt acatttacct
     1141 aacttaaacg aagaacaacg aaacgccttc atccaaagtt taaaagatga cccaagccaa
     1201 agcgctaacc ttttagcaga agctaaaaag ctaaatggtg ctcaggcgcc gaaagtagac
     1261 gcgaactcgg aattcaccat ggaacagaaa ctgatctctg aagaagacct ggtcttcaca
     1321 ctcgaagatt tcgttgggga ctggcgacag acagccggct acaacctgga ccaagtcctt
     1381 gaacagggag gtgtgtccag tttgtttcag aatctcgggg tgtccgtaac tccgatccaa
     1441 aggattgtcc tgagcggtga aaatgggctg aagatcgaca tccatgtcat catcccgtat
     1501 gaaggtctga gcggcgacca aatgggccag atcgaaaaaa tttttaaggt ggtgtaccct
     1561 gtggatgatc atcactttaa ggtgatcctg cactatggca cactggtaat cgacggggtt
     1621 acgccgaaca tgatcgacta tttcggacgg ccgtatgaag gcatcgccgt gttcgacggc
     1681 aaaaagatca ctgtaacagg gaccctgtgg aacggcaaca aaattatcga cgagcgcctg
     1741 atcaaccccg acggctccct gctgttccga gtaaccatca acggagtgac cggctggcgg
     1801 ctgtgcgaac gcattctggc gtgaaagctt ggtaccgagc tcggatccac tagtccagtg
     1861 tggtggaatt ctgcagatat ccagcacagt ggcggccgct cgagtctaga gggcccgttt
     1921 aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     1981 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     2041 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     2101 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     2161 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     2221 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2281 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2341 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     2401 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     2461 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     2521 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     2581 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     2641 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     2701 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     2761 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     2821 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     2881 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     2941 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     3001 tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3061 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3121 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     3181 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     3241 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     3301 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     3361 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     3421 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     3481 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     3541 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     3601 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     3661 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     3721 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     3781 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     3841 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     3901 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     3961 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     4021 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     4081 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     4141 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     4201 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     4261 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     4321 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     4381 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4441 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4501 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4561 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4621 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4681 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4741 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4801 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4861 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4921 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4981 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5041 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5101 ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca agcagcagat tacgcgcaga
     5161 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5221 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5281 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5341 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5401 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5461 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5521 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5581 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5641 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5701 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5761 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5821 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5881 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5941 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6001 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6061 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6121 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6181 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6241 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6301 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
