LOCUS       sequence-437765-        5470 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mScarlet in cells with TDP-43 loss-of-function. Leaky but 
            very sensitive to mild TDP-43 loss of function. It uses a cryptic 
            exon. [Code 'B5'] Note: It is not recommended to produce 
            lentiviruses containing TDP-REG sequences – see Depositor Comments.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5470)
  AUTHORS   Wilkins OG, Chien MZYJ, Wlaschin JJ, Barattucci S, Harley P, Mattedi
            F, Mehta PR, Pisliakova M, Ryadnov E, Keuss MJ, Thompson D, Digby H,
            Knez L, Simkin RL, Diaz JA, Zanovello M, Brown AL, Darbey A, Karda 
            R, Fisher EMC, Cunningham TJ, Le Pichon CE, Ule J, Fratta P
  TITLE     Creation of de novo cryptic splicing for ALS and FTD precision 
            medicine.
  JOURNAL   Science. 2024 Oct 4;386(6717):61-69. doi: 10.1126/science.adk2539. 
            Epub 2024 Oct 3.
  PUBMED    39361759
REFERENCE   2  (bases 1 to 5470)
  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: 
            "10.1126/science.adk2539"; journalName: "Science"; date: "2024-10-4-
            4"; volume: "386"; issue: "6717"; pages: "61-69"
COMMENT     Sequence Label: TDP-REGv2(mScarlet-FLAG)#10 in pTwist-CMV back...
FEATURES             Location/Qualifiers
     source          1..5470
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        16..319
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        321..523
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     474..494
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      628..637
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1611..1634
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1932..2053
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1943..1962)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1997..2016
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2168..2214
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2244..2380
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2425..2496
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2722..3310)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2802..2821)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(4056..4124,4125..4916)
                     /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     complement(4274..4293)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      4929..5065
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5217..5288
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5293..5339)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5357..5405
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
       61 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      121 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      181 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      241 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      301 agtcatcgct attaccatgc tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      361 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      421 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      481 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      541 gatcagatct ttgtcgatcc taccatccac tcgacacacc cgccagcggc cgcttcttgg
      601 tgccagctta tcatagcgct accggtcgcc accatggcga gaacaatggt cgcgatggta
      661 tctaagggcg aagcaggtaa gcggcgtgct tgttgcgtgg ttggggtgtg ggtgtgagtg
      721 ggatgggaga gtggttgtcg cgtgtggttg gctcgggtgc ttggatgggt gattgtcggc
      781 gtgtttgaca gtgataaaag agtttatgag attcaaagtc cacatggagg gatcaatgaa
      841 cggacacgaa tttgaaattg aaggcgaggg cgaaggaaga ccttatgagg ggacacagac
      901 cgccaaggtg cgtgcgtgga tcgtgtgcat gtggggtggt tgattagggg tgtatggctg
      961 ggtgattgag gcgtgtatgg tggtgtggat gacaagagtg attgttggtg tgaatgacga
     1021 gtgactgtct aacgtcttga ccgattctac agttgaaggt tacgaagggc gggcctctgc
     1081 ccttctcttg ggatatcctg agcccgcagt ttatgtacgg cagccgggct ttcaccaaac
     1141 accctgccga tatcccagac tactataaac agtcctttcc agaaggattt aagtgggagc
     1201 gagtcatgaa tttcgaggac ggaggtgccg tgacggttac tcaggacacc agcctggagg
     1261 acggcaccct gatctacaag gtgaagctga ggggcaccaa cttccccccc gacggccccg
     1321 tgatgcagaa gaagaccatg ggctgggagg ccagcaccga gaggctgtac cccgaggacg
     1381 gcgtgctgaa gggcgacatc aagatggccc tgaggctgaa ggacggcggc aggtacctgg
     1441 ccgacttcaa gaccacctac aaggccaaga agcccgtgca gatgcccggc gcctacaacg
     1501 tggacaggaa gctggacatc accagccaca acgaggacta caccgtggtg gagcagtacg
     1561 agaggagcga gggcaggcac agcaccggcg gcatggacga gctgtacaag gactacaagg
     1621 acgatgatga caagtgataa acaaatggta aggaagggca catcaatctt tgcttaattg
     1681 tcctttactc taaagatgta ttttatcata ctgaatgcta aacttgatat ctccttttag
     1741 gtcattgatg tccttcaccc cgggaaggcg acagtgccta agacagaaat tcgggaaaaa
     1801 ctagccaaaa tgtacaagac cacaccggat gtcatctttg tatttggatt cagaactcag
     1861 taaactggat ccgcaggcct ctgctagctt gactgactga gatacagcgt accttcagct
     1921 cacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     1981 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     2041 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt
     2101 gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtattggcc catctctatc
     2161 ggtatcgtag cataacccct tggggcctct aaacgggtct tgaggggttt tttgtgcccc
     2221 tcgggccgga ttgctatcta ccggcattgg cgcagaaaaa aatgcctgat gcgacgctgc
     2281 gcgtcttata ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc
     2341 acttccatac gtgtcctcct taccagaaat ttatccttaa ggtcgtcagc tatcctgcag
     2401 gcgatctctc gatttcgatc aagacattcc tttaatggtc ttttctggac accactaggg
     2461 gtcagaagta gttcatcaaa ctttcttccc tccctaatct cattggttac cttgggctat
     2521 cgaaacttaa ttaaccagtc aagtcagcta cttggcgaga tcgacttgtc tgggtttcga
     2581 ctacgctcag aattgcgtca gtcaagttcg atctggtcct tgctattgca cccgttctcc
     2641 gattacgagt ttcatttaaa tcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     2701 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     2761 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     2821 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     2881 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     2941 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3001 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3061 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3121 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     3181 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3241 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3301 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3361 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3421 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3481 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3541 agttgcattt aaatttccga actctccaag gccctcgtcg gaaaatcttc aaacctttcg
     3601 tccgatccat cttgcaggct acctctcgaa cgaactatcg caagtctctt ggccggcctt
     3661 gcgccttggc tattgcttgg cagcgcctat cgccaggtat tactccaatc ccgaatatcc
     3721 gagatcggga tcacccgaga gaagttcaac ctacatcctc aatcccgatc tatccgagat
     3781 ccgaggaata tcgaaatcgg ggcgcgcctg gtgtaccgag aacgatcctc tcagtgcgag
     3841 tctcgacgat ccatatcgtt gcttggcagt cagccagtcg gaatccagct tgggacccag
     3901 gaagtccaat cgtcagatat tgtactcaag cctggtcacg gcagcgtacc gatctgttta
     3961 aacctagata ttgatagtct gatcggtcaa cgtataatcg agtcctagct tttgcaaaca
     4021 tctatcaaga gacaggatca gcaggaggct ttcgcatgag tattcaacat ttccgtgtcg
     4081 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     4141 tgaaagtaaa agatgctgaa gatcagttgg gtgcgcgagt gggttacatc gaactggatc
     4201 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgctttcca atgatgagca
     4261 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     4321 tcggtcgccg catacactat tctcagaatg acttggttga gtattcacca gtcacagaaa
     4381 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     4441 ataacactgc ggccaactta cttctgacaa cgattggagg accgaaggag ctaaccgctt
     4501 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     4561 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaaccttgc
     4621 gtaaactatt aactggcgaa ctacttactc tagcttcccg gcaacagttg atagactgga
     4681 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     4741 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     4801 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     4861 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaaccga
     4921 ttctaggtgc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc
     4981 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc
     5041 ctccttacca gaaatttatc cttaagatcc cgaatcgttt aaactcgact ctggctctat
     5101 cgaatctccg tcgtttcgag cttacgcgaa cagccgtggc gctcatttgc tcgtcgggca
     5161 tcgaatctcg tcagctatcg tcagcttacc tttttggcag cgatcgcggc tcccgacatc
     5221 ttggaccatt agctccacag gtatcttctt ccctctagtg gtcataacag cagcttcagc
     5281 tacctctcaa ttcaaaaaac ccctcaagac ccgtttagag gccccaaggg gttatgctat
     5341 caatcgttgc gttacacaca caaaaaacca acacacatcc atcttcgatg gatagcgatt
     5401 ttattatcta actgctgatc gagtgtagcc agatctagta atcaattacg gggtcattag
     5461 ttcatagccc
//
