LOCUS       sequence-324209-        5817 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows for transcription of mVenus fluorophore half fused to TDP-43 
            for injection into zebrafish embryos for BiFC assays.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5817)
  AUTHORS   Don EK, Maschirow A, Radford RAW, Scherer NM, Vidal-Itriago A, Hogan
            A, Maurel C, Formella I, Stoddart JJ, Hall TE, Lee A, Shi B, Cole 
            NJ, Laird AS, Badrock AP, Chung RS, Morsch M
  TITLE     In vivo Validation of Bimolecular Fluorescence Complementation 
            (BiFC) to Investigate Aggregate Formation in Amyotrophic Lateral 
            Sclerosis (ALS).
  JOURNAL   Mol Neurobiol. 2021 May;58(5):2061-2074. doi: 
            10.1007/s12035-020-02238-0. Epub 2021 Jan 7.
  PUBMED    33415684
REFERENCE   2  (bases 1 to 5817)
  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: 
            "10.1007/s12035-020-02238-0"; journalName: "Mol Neurobiol"; date: 
            "2021-05"; volume: "58"; issue: "5"; pages: "2061-2074"
COMMENT     Sequence Label: pCS2plus-mVenus1-155-GGGS-TDP-43
FEATURES             Location/Qualifiers
     source          1..5817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      88..97
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(94..96,97..99,100..558)
                     /codon_start=1
                     /product="improved N-terminal fragment of mVenus for use in
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN155(I152L)
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITA"
     primer_bind     complement(139..160)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(400..419)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    1869..2003
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1893..1912)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1947..1966
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2116..2136)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2116..2134)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2155..2171)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2155..2171)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2168..2190)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2179..2195
                     /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(2203..2209,2210..2227,2228..2233))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2248..2269
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2386..2403)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2557..3145)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2637..2656)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3316..4107,4108..4176))
                     /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     3939..3958
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4177..4281)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4307..4762)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4444..4465)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4656..4675
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4833..5817
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atccgccgcc accatggtga gcaagggcga ggagctgttc
      121 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      181 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctgatctgc
      241 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctggg ctacggcctg
      301 cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
      361 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
      421 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
      481 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
      541 aacgtctata tcaccgccgg cggaggaagc ggaggcggaa gcggcggggg aagcatgtct
      601 gaatatattc gggtaaccga agatgagaac gatgagccca ttgaaatacc atcggaagac
      661 gatgggacgg tgctgctctc cacggttaca gcccagtttc caggggcgtg tgggcttcgc
      721 tacaggaatc cagtgtctca gtgtatgaga ggtgtccggc tggtagaagg aattctgcat
      781 gccccagatg ctggctgggg aaatctggtg tatgttgtca actatccaaa agataacaaa
      841 agaaaaatgg atgagacaga tgcttcatca gcagtgaaag tgaaaagagc agtccagaaa
      901 acatccgatt taatagtgtt gggtctccca tggaaaacaa ccgaacagga cctgaaagag
      961 tattttagta cctttggaga agttcttatg gtgcaggtca agaaagatct taagactggt
     1021 cattcaaagg ggtttggctt tgttcgtttt acggaatatg aaacacaagt gaaagtaatg
     1081 tcacagcgac atatgataga tggacgatgg tgtgactgca aacttcctaa ttctaagcaa
     1141 agccaagatg agcctttgag aagcagaaaa gtgtttgtgg ggcgctgtac agaggacatg
     1201 actgaggatg agctgcggga gttcttctct cagtacgggg atgtgatgga tgtcttcatc
     1261 cccaagccat tcagggcctt tgcctttgtt acatttgcag atgatcagat tgcgcagtct
     1321 ctttgtggag aggacttgat cattaaagga atcagcgttc atatatccaa tgccgaacct
     1381 aagcacaata gcaatagaca gttagaaaga agtggaagat ttggtggtaa tccaggtggc
     1441 tttgggaatc agggtggatt tggtaatagc agagggggtg gagctggttt gggaaacaat
     1501 caaggtagta atatgggtgg tgggatgaac tttggtgcgt tcagcattaa tccagccatg
     1561 atggctgccg cccaggcagc actacagagc agttggggta tgatgggcat gttagccagc
     1621 cagcagaacc agtcaggccc atcgggtaat aaccaaaacc aaggcaacat gcagagggag
     1681 ccaaaccagg ccttcggttc tggaaataac tcttatagtg gctctaattc tggtgcagca
     1741 attggttggg gatcagcatc caatgcaggg tcgggcagtg gttttaatgg aggctttggc
     1801 tcaagcatgg attctaagtc ttctggctgg ggaatgtagt ctagaactat agtgagtcgt
     1861 attacgtaga tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     1921 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     1981 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     2041 agggggaggt gtgggaggtt ttttaattcg cggccgcggc gccaatgcat tgggcccggt
     2101 acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata
     2161 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccgggag
     2221 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     2281 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     2341 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     2401 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     2461 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     2521 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2581 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2641 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2701 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2761 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2821 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2881 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2941 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3001 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3061 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3121 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3181 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3241 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3301 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     3361 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     3421 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     3481 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3541 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3601 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3661 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3721 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3781 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3841 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3901 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3961 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4021 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4081 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     4141 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     4201 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     4261 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     4321 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     4381 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     4441 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     4501 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     4561 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     4621 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     4681 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     4741 aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg
     4801 cgatcggtgc gggcctcttc gctattacgc cagtcgaccg ccaattcaat atggcgtata
     4861 tggactcatg ccaattcaat atggtggatc tggacctgtg ccaattcaat atggcgtata
     4921 tggactcgtg ccaattcaat atggtggatc tggaccccag ccaattcaat atggcggact
     4981 tggcaccatg ccaattcaat atggcggacc tggcactgtg ccaactgggg aggggtctac
     5041 ttggcacggt gccaagtttg aggaggggtc ttggccctgt gccaagtccg ccatattgaa
     5101 ttggcatggt gccaataatg gcggccatat tggctatatg ccaggatcaa tatataggca
     5161 atatccaata tggccctatg ccaatatggc tattggccag gttcaatact atgtattggc
     5221 cctatgccat atagtattcc atatatgggt tttcctattg acgtagatag cccctcccaa
     5281 tgggcggtcc catataccat atatggggct tcctaatacc gcccatagcc actcccccat
     5341 tgacgtcaat ggtctctata tatggtcttt cctattgacg tcatatgggc ggtcctattg
     5401 acgtatatgg cgcctccccc attgacgtca attacggtaa atggcccgcc tggctcaatg
     5461 cccattgacg tcaataggac cacccaccat tgacgtcaat gggatggctc attgcccatt
     5521 catatccgtt ctcacgcccc ctattgacgt caatgacggt aaatggccca cttggcagta
     5581 catcaatatc tattaatagt aacttggcaa gtacattact attggaagta cgccagggta
     5641 cattggcagt actcccattg acgtcaatgg cggtaaatgg cccgcgatgg ctgccaagta
     5701 catccccatt gacgtcaatg gggaggggca atgacgcaaa tgggcgttcc attgacgtaa
     5761 atgggcggta ggcgtgccta atgggaggtc tatataagca atgctcgttt agggaac
//
