LOCUS       sequence-437763-        5358 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mScarlet in cells with TDP-43 loss-of-function. Very 
            bright and decent dynamic range (>10-fold), slightly leaky. It uses 
            a single intron. [Code 'B12']. 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 5358)
  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 5358)
  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)#11 in pTwist-CMV back...
FEATURES             Location/Qualifiers
     source          1..5358
                     /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             1499..1522
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1820..1941
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1831..1850)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1885..1904
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2056..2102
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2132..2268
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2313..2384
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2610..3198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2690..2709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(3944..4012,4013..4804)
                     /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(4162..4181)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      4817..4953
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5105..5176
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5181..5227)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5245..5293
                     /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 tgctatggtg
      661 tccaaaggtg aggcagtcat aaaggagttt atgaggttca aggtgcacat ggaagggtca
      721 atgaacggac atgagttcga gatcgaaggt gaaggagagg gccgacccta cgaagggacg
      781 cagacggcca agctgaaggt gacgaaaggc ggaccgttgc cattctcgtg ggatattctg
      841 tcacctcagg taagtatcgg tttttgttgt ttgcaagagt gaaaaagtgt agaaaactaa
      901 attattagat gtcgggtggt gctgctcgtt agtctaactc tagccgttaa ctttgtgtgt
      961 gtgtgtgtgt gtgaatgtgt gtgtgtgtgt gtgttagttc atgtacggca gtcgtgcgtt
     1021 cacgaaacac cctgctgaca tccctgacta ctacaaacag tcctttcccg agggcttcaa
     1081 atgggaacgt gtcatgaatt ttgaagacgg aggggccgtg accgtcactc aagatacgtc
     1141 gttggaagac ggaacgttga tatataaggt caaattgcgc ggaacaaact tcccacccga
     1201 tggtccagta atgcaaaaga agactatggg ttgggaggca agtacagaga ggctgtaccc
     1261 ggaagatggt gttctaaaag gagatatcaa gatggccctg aggcttaagg acggtgggcg
     1321 atacctggcc gacttcaaga ccacctacaa ggccaagaag cccgtgcaga tgcccggcgc
     1381 ctacaacgtg gacaggaagc tggacatcac cagccacaac gaggactaca ccgtggtgga
     1441 gcagtacgag aggagcgagg gcaggcacag caccggcggc atggacgagc tgtacaagga
     1501 ctacaaggac gatgatgaca agtgataaac aaatggtaag gaagggcaca tcaatctttg
     1561 cttaattgtc ctttactcta aagatgtatt ttatcatact gaatgctaaa cttgatatct
     1621 ccttttaggt cattgatgtc cttcaccccg ggaaggcgac agtgcctaag acagaaattc
     1681 gggaaaaact agccaaaatg tacaagacca caccggatgt catctttgta tttggattca
     1741 gaactcagta aactggatcc gcaggcctct gctagcttga ctgactgaga tacagcgtac
     1801 cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     1861 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     1921 ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag
     1981 ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tattggccca
     2041 tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     2101 tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa tgcctgatgc
     2161 gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca
     2221 acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg tcgtcagcta
     2281 tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt ttctggacac
     2341 cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca ttggttacct
     2401 tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc gacttgtctg
     2461 ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg ctattgcacc
     2521 cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca aaaggccagg
     2581 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2641 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2701 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2761 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2821 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2881 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2941 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     3001 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     3061 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     3121 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     3181 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     3241 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     3301 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     3361 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3421 tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga aaatcttcaa
     3481 acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca agtctcttgg
     3541 ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta ctccaatccc
     3601 gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa tcccgatcta
     3661 tccgagatcc gaggaatatc gaaatcgggg cgcgcctggt gtaccgagaa cgatcctctc
     3721 agtgcgagtc tcgacgatcc atatcgttgc ttggcagtca gccagtcgga atccagcttg
     3781 ggacccagga agtccaatcg tcagatattg tactcaagcc tggtcacggc agcgtaccga
     3841 tctgtttaaa cctagatatt gatagtctga tcggtcaacg tataatcgag tcctagcttt
     3901 tgcaaacatc tatcaagaga caggatcagc aggaggcttt cgcatgagta ttcaacattt
     3961 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     4021 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcgcgagtgg gttacatcga
     4081 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gctttccaat
     4141 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     4201 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt attcaccagt
     4261 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     4321 catgagtgat aacactgcgg ccaacttact tctgacaacg attggaggac cgaaggagct
     4381 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     4441 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     4501 aaccttgcgt aaactattaa ctggcgaact acttactcta gcttcccggc aacagttgat
     4561 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     4621 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     4681 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     4741 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     4801 gtaaccgatt ctaggtgcat tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt
     4861 atactcccac atatgccaga ttcagcaacg gatacggctt ccccaacttg cccacttcca
     4921 tacgtgtcct ccttaccaga aatttatcct taagatcccg aatcgtttaa actcgactct
     4981 ggctctatcg aatctccgtc gtttcgagct tacgcgaaca gccgtggcgc tcatttgctc
     5041 gtcgggcatc gaatctcgtc agctatcgtc agcttacctt tttggcagcg atcgcggctc
     5101 ccgacatctt ggaccattag ctccacaggt atcttcttcc ctctagtggt cataacagca
     5161 gcttcagcta cctctcaatt caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt
     5221 tatgctatca atcgttgcgt tacacacaca aaaaaccaac acacatccat cttcgatgga
     5281 tagcgatttt attatctaac tgctgatcga gtgtagccag atctagtaat caattacggg
     5341 gtcattagtt catagccc
//
