LOCUS       sequence-324218-        5817 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows for transcription of improved mVenus I152L fluorophore half 
            fused to mutant TDP-43-M337V 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-I152L-GGGS-TDP43 M337V
FEATURES             Location/Qualifiers
     source          1..5817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        804..822
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     804..821
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      857..866
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(863..865,866..868,869..1327)
                     /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
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYLTA"
     primer_bind     complement(908..929)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1169..1188)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2638..2772
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2662..2681)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2716..2735
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2885..2905)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2885..2903)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2924..2940)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2924..2940)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2937..2959)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2948..2964
                     /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(2972..2978,2979..2996,2997..3002))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3017..3038
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3155..3172)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3326..3914)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3406..3425)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4085..4876,4877..4945))
                     /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     4708..4727
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4946..5050)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5076..5531)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5213..5234)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5425..5444
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5602..5817,1..769)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 gtgccaagtt tgaggagggg tcttggccct gtgccaagtc cgccatattg aattggcatg
       61 gtgccaataa tggcggccat attggctata tgccaggatc aatatatagg caatatccaa
      121 tatggcccta tgccaatatg gctattggcc aggttcaata ctatgtattg gccctatgcc
      181 atatagtatt ccatatatgg gttttcctat tgacgtagat agcccctccc aatgggcggt
      241 cccatatacc atatatgggg cttcctaata ccgcccatag ccactccccc attgacgtca
      301 atggtctcta tatatggtct ttcctattga cgtcatatgg gcggtcctat tgacgtatat
      361 ggcgcctccc ccattgacgt caattacggt aaatggcccg cctggctcaa tgcccattga
      421 cgtcaatagg accacccacc attgacgtca atgggatggc tcattgccca ttcatatccg
      481 ttctcacgcc ccctattgac gtcaatgacg gtaaatggcc cacttggcag tacatcaata
      541 tctattaata gtaacttggc aagtacatta ctattggaag tacgccaggg tacattggca
      601 gtactcccat tgacgtcaat ggcggtaaat ggcccgcgat ggctgccaag tacatcccca
      661 ttgacgtcaa tggggagggg caatgacgca aatgggcgtt ccattgacgt aaatgggcgg
      721 taggcgtgcc taatgggagg tctatataag caatgctcgt ttagggaacc gccattctgc
      781 ctggggacgt cggagcaagc ttgatttagg tgacactata gaatacaagc tacttgttct
      841 ttttgcagga tccgccgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
      901 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
      961 gggcgagggc gatgccacct acggcaagct gaccctgaag ctgatctgca ccaccggcaa
     1021 gctgcccgtg ccctggccca ccctcgtgac caccctgggc tacggcctgc agtgcttcgc
     1081 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     1141 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     1201 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     1261 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatct
     1321 caccgccggc ggaggaagcg gaggcggaag cggcggggga agcatgtctg aatatattcg
     1381 ggtaaccgaa gatgagaacg atgagcccat tgaaatacca tcggaagacg atgggacggt
     1441 gctgctctcc acggttacag cccagtttcc aggggcgtgt gggcttcgct acaggaatcc
     1501 agtgtctcag tgtatgagag gtgtccggct ggtagaagga attctgcatg ccccagatgc
     1561 tggctgggga aatctggtgt atgttgtcaa ctatccaaaa gataacaaaa gaaaaatgga
     1621 tgagacagat gcttcatcag cagtgaaagt gaaaagagca gtccagaaaa catccgattt
     1681 aatagtgttg ggtctcccat ggaaaacaac cgaacaggac ctgaaagagt attttagtac
     1741 ctttggagaa gttcttatgg tgcaggtcaa gaaagatctt aagactggtc attcaaaggg
     1801 gtttggcttt gttcgtttta cggaatatga aacacaagtg aaagtaatgt cacagcgaca
     1861 tatgatagat ggacgatggt gtgactgcaa acttcctaat tctaagcaaa gccaagatga
     1921 gcctttgaga agcagaaaag tgtttgtggg gcgctgtaca gaggacatga ctgaggatga
     1981 gctgcgggag ttcttctctc agtacgggga tgtgatggat gtcttcatcc ccaagccatt
     2041 cagggccttt gcctttgtta catttgcaga tgatcagatt gcgcagtctc tttgtggaga
     2101 ggacttgatc attaaaggaa tcagcgttca tatatccaat gccgaaccta agcacaatag
     2161 caatagacag ttagaaagaa gtggaagatt tggtggtaat ccaggtggct ttgggaatca
     2221 gggtggattt ggtaatagca gagggggtgg agctggtttg ggaaacaatc aaggtagtaa
     2281 tatgggtggt gggatgaact ttggtgcgtt cagcattaat ccagccatga tggctgccgc
     2341 ccaggcagca ctacagagca gttggggtat ggtgggcatg ttagccagcc agcagaacca
     2401 gtcaggccca tcgggtaata accaaaacca aggcaacatg cagagggagc caaaccaggc
     2461 cttcggttct ggaaataact cttatagtgg ctctaattct ggtgcagcaa ttggttgggg
     2521 atcagcatcc aatgcagggt cgggcagtgg ttttaatgga ggctttggct caagcatgga
     2581 ttctaagtct tctggctggg gaatgtagtc tagaactata gtgagtcgta ttacgtagat
     2641 ccagacatga taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa
     2701 aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc
     2761 aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg
     2821 tgggaggttt tttaattcgc ggccgcggcg ccaatgcatt gggcccggta cccagctttt
     2881 gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg
     2941 tgtgaaattg ttatccgctc acaattccac acaacatacg agccgggagc ataaagtgta
     3001 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3061 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     3121 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3181 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3241 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3301 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3361 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3421 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3481 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3541 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3601 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3661 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3721 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3781 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3841 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3901 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3961 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4021 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4081 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     4141 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     4201 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     4261 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     4321 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     4381 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4441 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4501 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4561 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4621 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4681 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4741 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4801 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4861 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4921 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4981 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     5041 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
     5101 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     5161 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     5221 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     5281 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     5341 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     5401 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     5461 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     5521 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     5581 ggcctcttcg ctattacgcc agtcgaccgc caattcaata tggcgtatat ggactcatgc
     5641 caattcaata tggtggatct ggacctgtgc caattcaata tggcgtatat ggactcgtgc
     5701 caattcaata tggtggatct ggaccccagc caattcaata tggcggactt ggcaccatgc
     5761 caattcaata tggcggacct ggcactgtgc caactgggga ggggtctact tggcacg
//
