LOCUS       sequence-395362-        5801 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NanoBit detection of V5-tagged protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5801)
  AUTHORS   Zeghal M, Matte K, Venes A, Patel S, Laroche G, Sarvan S, Joshi M, 
            Rain JC, Couture JF, Giguere PM
  TITLE     Development of a V5-tag-directed nanobody and its implementation as 
            an intracellular biosensor of GPCR signalling.
  JOURNAL   J Biol Chem. 2023 Jul 28:105107. doi: 10.1016/j.jbc.2023.105107.
  PUBMED    37517699
REFERENCE   2  (bases 1 to 5801)
  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: "J Biol 
            Chem. 2023 Jul 28:105107. doi: 10.1016/j.jbc.2023.105107."
COMMENT     Sequence Label: NbV5-HiBiT
FEATURES             Location/Qualifiers
     source          1..5801
                     /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"
     primer_bind     complement(1392..1409)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1398..1622
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1668..2096
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1755..1774)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1965..1986
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2105..2125)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2110..2439
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2290..2425
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2352..2371
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2506..3300
                     /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(2560..2579)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3170..3189
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3474..3595
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3511..3530)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3565..3584
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3644..3660)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3644..3660)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3657..3679)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3668..3684
                     /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(3692..3698,3699..3716,3717..3722))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3737..3758
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3875..3892)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4046..4634)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4126..4145)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4805..5596,5597..5665))
                     /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     5428..5447
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5666..5770)
                     /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 gtttaaactt aagcttacca tggctgaagt gcagctgcaa gccagcggcg gaggcttcgt
      961 gcagcccggc ggatctctga gactgtcttg tgccgcttct ggcaccacca gcttcggcga
     1021 tacaatgggc tggttccggc aggcccctgg aaaggaaaga gagtttgtgt ccgccatctc
     1081 cagacagggc gacaagcact actatgccga cagcgtgaaa ggcagattca caatcagccg
     1141 ggacaacagc aagaacaccg tgtacctgca gatgaactct ctgagagccg aagataccgc
     1201 cacctactac tgcgccgagt ggatgaatac cagacgcgag ttcatcaccc cttactgggg
     1261 ccagggaaca caggtgaccg tgtctagcgg atccggtggt ggcgggagcg gaggtggagg
     1321 ctcggtgagc ggctggcggc tgttcaagaa gattagctaa tctagagggc ccgtttaaac
     1381 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     1441 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     1501 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     1561 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     1621 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     1681 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     1741 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     1801 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     1861 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     1921 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     1981 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     2041 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     2101 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     2161 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     2221 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     2281 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     2341 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     2401 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     2461 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     2521 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     2581 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     2641 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     2701 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     2761 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     2821 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     2881 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     2941 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     3001 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     3061 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     3121 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     3181 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     3241 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     3301 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     3361 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     3421 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     3481 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     3541 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     3601 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     3661 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     3721 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3781 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     3841 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3901 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3961 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4021 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4081 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4141 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4201 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4261 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4321 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4381 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4441 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     4501 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4561 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4621 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4681 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4741 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4801 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     4861 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     4921 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     4981 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5041 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5101 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5161 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5221 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5281 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5341 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5401 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5461 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5521 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5581 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5641 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     5701 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     5761 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
