LOCUS       sequence-380387-        7135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express DjCas13d in a Cre dependent manner.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7135)
  AUTHORS   Back F, Sandoval A, Vu LM, Hong VM, Bhaskara A, Rodriguez SR, 
            O'Brien JT, Kolber BJ, Kroener S, Ploski JE
  TITLE     Adeno-associated viral vector resource for the RNA-targeting Cas13d:
            A comparison of high-fidelity variants, DjCas13d and hfCas13d.
  JOURNAL   Mol Ther Methods Clin Dev. 2025 Aug 20;33(4):101565. doi: 
            10.1016/j.omtm.2025.101565. eCollection 2025 Dec 11.
  PUBMED    41020266
REFERENCE   2  (bases 1 to 7135)
  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: "Mol Ther 
            Methods Clin Dev."; date: "2025-08-20"; volume: "33(4):101565. doi";
            pages: " 10.1016/j.omtm.2025.101565. eCollection 2025 Dec 11"
COMMENT     Sequence Label: pAAV-CMV-intron-DIO-DjCas13d-pA
FEATURES             Location/Qualifiers
     source          1..7135
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      22..610
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     511..530
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   672..801
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        881..1184
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1185..1388
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1339..1359
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1385..1409
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          1537..1915
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
     protein_bind    complement(1967..2000)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(2051..2084)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2096..2122)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2132..2152)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(4802..4822)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    4841..4874
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    4925..4958
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5011..5030)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5069..5209
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5069..5198
                     /label=AAV2 ITR
     rep_origin      5284..5739
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5371..5390)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5581..5602
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5756..5775)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5875..5897
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5935..5953)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6021..6125
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6126..6194,6195..6986)
                     /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(6344..6363)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
       61 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
      121 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
      181 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
      241 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
      301 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
      361 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
      421 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
      481 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
      541 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
      601 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
      661 tgctcacatg tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc
      721 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      781 actccatcac taggggttcc tgcggccgca cgcgtggagc tagttattaa tagtaatcaa
      841 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      901 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      961 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     1021 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
     1081 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
     1141 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     1201 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
     1261 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
     1321 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
     1381 gcagagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgttttgacc
     1441 tccatagaag acaccgggac cgatccagcc tccgcggatt cgaatcccgg ccgggaacgg
     1501 tgcattggaa cgcggattcc ccgtgccaag agtgacgtaa gtaccgccta tagagtctat
     1561 aggcccacaa aaaatgcttt cttcttttaa tatacttttt tgtttatctt atttctaata
     1621 ctttccctaa tctctttctt tcagggcaat aatgatacaa tgtatcatgc ctctttgcac
     1681 cattctaaag aataacagtg ataatttctg ggttaaggca atagcaatat ttctgcatat
     1741 aaatatttct gcatataaat tgtaactgat gtaagaggtt tcatattgct aatagcagct
     1801 acaatccagc taccattctg cttttatttt atggttggga taaggctgga ttattctgag
     1861 tccaagctag gcccttttgc taatcatgtt catacctctt atcttcctcc cacagctcct
     1921 gggcaacgtg ctggtctgtg tgctggccca tcacgtcgac tccggaataa cttcgtatag
     1981 gatactttat acgaagttat gcagaatggt agctggattg tagctgctat tagcaatatg
     2041 aaacctctta ataacttcgt atagcataca ttatacgaag ttatggcgcg ccctaagcgt
     2101 aatctggaac atcgtatggg taagcggccg ccaccttcct ctttttctta ggtccggatc
     2161 ccacttcctc ttttggttcg ggattcgggt tatttcgatc aaaaagttca tttatcgtga
     2221 ggtttttgta ccgcggcacg acgtatccga acgggataca gtacgctttt acaaagtcct
     2281 tacaataagt gtggtgcttt ttcagattgt tcagatattc ggcggtcttc ggctggtggc
     2341 tggtatcata ccaattattc tttgcgatca ttgattgaac gagatagtga taaagttcga
     2401 agtaagagtg cagttctccg atcccgctta tgtattcatt actattcctc acaacggata
     2461 tgtgagcaac tatgttgcgg aatgaactca cagctgttat gtcgctattc tgaaggttat
     2521 ttttgacaag aacgtaccat ttgcggtgtc ctttctggtt cttcaggtgg ccgtacgtag
     2581 tgtaattttt gtcagtcaag aggtgagtag tcaattgact gtacacaatt tttttattca
     2641 tcttagggag cgtcacaccg tgaaggaatg cgtctcgttc caggcaatga aatgcaatca
     2701 catacctact attgacatac accaggtttt tggttataag gtacataaca gtcaaataaa
     2761 gacttacaat tgcctggtac cgctgttttt tcagcgcgtc actgaagttt cggctaggat
     2821 catctttatt gatctttgca ctttggggta cgtcttcaaa tattttatag tctactgaac
     2881 taattattcg ggcaagaacg tctatctgtg cggacttgtc agacgtcggt gggatctctt
     2941 gacctttaca aacatcaaag tatcgtttta tttgtgtctc cggaatccct tcgagcacaa
     3001 attttactac attagggttt ttcacaagag tccgtgtctt gtgtggatta ttataccgta
     3061 cgaggtaatg gaagcgtgaa gattcaataa cgttggaagc aatgaaattg cgaaagccct
     3121 gtttaccttt gataaatttg ccatccgcac ccacgggcat tgtcctttcg agttcccgat
     3181 caatctcttc atccggccga ttgtccattc ccaatatgcg cagagcatcg cgatacatga
     3241 tctttcgggc gtcggcgctt ggcttttgca ttctagcgat actgttgata acccgcagct
     3301 cttcgagagt atacctgcag ttggtcatgt taaagaagtc cgcgtattca ggcttgaatt
     3361 taaattccaa attgagtttc ttcaggatgt ctatgaagga gattatatta tcaaatttgt
     3421 tgatcagggt tgtgaggagg tcgtttatct cctttccatc gaggagaagg gtgagcatgt
     3481 atatgagctc tgagaaatac ccaatgcctg tgttccctat tttaatatct tcaaaactta
     3541 tcctatagtc gttcgccttc ttctgatagt cccttatgtt tttggcattt acatcggcag
     3601 cggctttctt gaaatcggcc ccaagagcgc tcgcgagtgt ccgggccgtt ttgatgtaaa
     3661 tctcttcctt gttttcctca gtgagtgaac ttcgcaagga gctcacgagc gcctccttct
     3721 caaaagcatg gtggtcgtag taataggtta taataaagtc gaagagctta tacagtttag
     3781 ccctgacgga gttgtactta tcctctttga aactcttaag atctgattgc tcgagcataa
     3841 tctctcgcaa tttcttgatg ctaaatccca tatttttaaa ggacttcttg atcaggaatt
     3901 cataataagc gcgtacaatc gtctccttac tctctttcgg gaaaacatga cacaaaatat
     3961 ggaggtttac tgtattagtt ttaaggaatt cactatctat cctttcggtg acctcttccc
     4021 acaaaatact gagagtctcc ttgaactcgt cagacagttg gtcctcgagt tgatacagcc
     4081 agaatgaatt cacgtggtcg ctctcatcac tactcagatc ggaatgaaag caaaactgtc
     4141 taagggcaga cagcagggta agtattgcga agatgtcttc ctcataccgt ctctcgtttt
     4201 catggtaaaa tgcctccccg aaataaagaa gttcttttct tttaatgtag tttttaaacc
     4261 tatcatattt gttagtctgc cctctaagag tttggtaatc gagattaatc cgccagcccc
     4321 ctgccccaat gaaatcttca gtcagctctt ccttttgcgg atgcaggaca ttgtccagtg
     4381 tgaatacgat gttgttaaca tgaaccgcaa ggatcttttc aatatccagt atttgatata
     4441 tcaattgtat gtggatattg tcgtcgaaag tcttcccaaa atagtgcatt tcgagtttgt
     4501 ctttcaggca cagaaggtcc atacctggct gcggcttaaa cagggggttg tcggtagtgg
     4561 cgatcttcgg gtgtcgacct tggaacttaa tctttttacc agatacttcg acgttcttca
     4621 agccggcgct actattgaga ttttggggct tttgagagct cgggatcttg taagagagat
     4681 tcgccatgtt ccccttccca aaagacgtca ttatgacagt cccctgcaca gaaagtgcct
     4741 ctttcaatcc ggaggtttta gaaaccgttt tcttggattt ttgcttcttg atgctggcct
     4801 ccacctttct cttcttcttg gggctcatgg tggcgctagc ataacttcgt ataaagtatc
     4861 ctatacgaag ttatttgcct taacccagaa attatcactg ttattcttta gaatggtgca
     4921 aagaataact tcgtataatg tatgctatac gaagttatga attcgatact agtctcgagt
     4981 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     5041 catttttttc actggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc
     5101 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     5161 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     5221 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     5281 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     5341 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5401 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5461 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     5521 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5581 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     5641 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     5701 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     5761 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     5821 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     5881 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     5941 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     6001 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     6061 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     6121 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     6181 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     6241 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     6301 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     6361 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     6421 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     6481 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     6541 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     6601 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     6661 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     6721 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     6781 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6841 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6901 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6961 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     7021 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     7081 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgt
//
