LOCUS       sequence-337347-        7934 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for encoding a human codon-optimized Cas13Y.1 
            driven by CMV promoter,mCherry driven by CMV promoter and U6-driven 
            crRNAs cloning site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7934)
  AUTHORS   Xu C, Zhou Y, Xiao Q, He B, Geng G, Wang Z, Cao B, Dong X, Bai W, 
            Wang Y, Wang X, Zhou D, Yuan T, Huo X, Lai J, Yang H
  TITLE     Programmable RNA editing with compact CRISPR-Cas13 systems from 
            uncultivated microbes.
  JOURNAL   Nat Methods. 2021 May;18(5):499-506. doi: 
            10.1038/s41592-021-01124-4. Epub 2021 May 3.
  PUBMED    33941935
REFERENCE   2  (bases 1 to 7934)
  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: 
            "10.1038/s41592-021-01124-4"; journalName: "Nat Methods"; date: 
            "2021-05"; volume: "18"; issue: "5"; pages: "499-506"
COMMENT     Sequence Label: CMV-Cas13Y.1-SV40pA_U6-BbsI-DR_CMV-mCherry-BGHpA
FEATURES             Location/Qualifiers
     source          1..7934
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   25..87
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        396..681
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        683..960
                     /label=chicken beta-actin promoter
     intron          961..1188
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1209..1229
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3597..3617
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3624..3689
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    3699..3833
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3736..3755)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3790..3809
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3840..4088
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     3840..3860
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4011..4030
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        4222..4525
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4526..4729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4680..4700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4758..4767
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4764..5474
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     DDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4909..4927)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     polyA_signal    5491..5715
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6072..6106
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        6137..6241
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6242..6310,6311..7102)
                     /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(6460..6479)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7273..7861
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7762..7781
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tatcccctga ttctctcgag ttaaccctag aaagataatc atattgtgac gtacgttaaa
       61 gataatcatg cgtaaaattg acgcatgtgt tttatcgatc tgtatatcga ggtttattta
      121 ttaatttgaa tagatattaa gttttattat atttacactt acatactaat aataaattca
      181 acaaacaatt tatttatgtt tatttattta ttaaaaaaaa acaaaaactc aaaatttctt
      241 ctataaagta acaaaacttt taaacattct ctcttttaca aaaataaact tattttgtac
      301 tttaaaaaca gtcatgttgt attataaaat aagtaattag cttaacttat acataataga
      361 aacaaattat acttattaat taatctagag gtacccgtta cataacttac ggtaaatggc
      421 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caatagtaac gccaataggg
      481 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      541 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      601 tggcattgtg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      661 ttagtcatcg ctattaccat ggtcgaggtg agccccacgt tctgcttcac tctccccatc
      721 tcccccccct ccccaccccc aattttgtat ttatttattt tttaattatt ttgtgcagcg
      781 atgggggcgg gggggggggg ggggcgcgcg ccrggsgggg sggggsgggg sgrggggsgg
      841 ggsggggsga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
      901 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
      961 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg
     1021 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct
     1081 ccgggctgta attagctgag caagaggtaa gggtttaagg gatggttggt tggtggggta
     1141 ttaatgttta attacctgga gcacctgcct gaaatcactt tttttcaggt tggaccggtg
     1201 ccaccatgcc caagaagaag cggaaggtga atggcatcga gctgaagaag gaagaagccg
     1261 ccttctactt caatcaggcc gagctgaacc tgaaggccat tgaggacaac atcttcgaca
     1321 aggagagacg gaaaacactg ctgaacaacc cccagatcct ggccaagatg gagaacttta
     1381 tcttcaattt ccgggacgtg accaagaacg ccaagggcga aatcgactgc ctgctgctga
     1441 agctgagaga gctgcggaac ttttacagcc actacgtgca caagcgggac gtcagagaac
     1501 tgagcaaggg cgagaagccg atcctggaga agtactacca gttcgccatc gaatccaccg
     1561 gctctgagaa cgtgaagctc gaaatcatcg aaaacgacgc ctggctggcc gacgccggcg
     1621 tgctgttctt cctgtgcatc ttcctgaaga agagccaggc aaacaagctg atcagcggca
     1681 tcagcggctt caagagaaac gacgacaccg gccagcctcg gagaaacctg ttcacctact
     1741 tctccatccg ggagggctac aaggtggtgc ccgaaatgca gaagcacttc ctgctgttct
     1801 ccctggtgaa ccacctgagc aaccaggacg attatatcga aaaggcccac cagccctacg
     1861 acatcggcga gggcctcttc ttccaccgga ttgccagcac cttcctgaac atctccggaa
     1921 tcctgagaaa catgaagttc tacacctatc agagcaagag actggtggag cagagaggcg
     1981 agctgaagcg ggaaaaggac atcttcgcct gggaagaacc gtttcagggc aattcctact
     2041 ttgagatcaa cggccacaag ggcgtgattg gcgaggacga gctgaaggag ctgtgctacg
     2101 ccttcctgat cggcaaccag gacgccaaca aggtggaggg ccggatcacc cagttcctgg
     2161 agaagttcag aaacgccaac agcgtgcagc aggtgaagga cgacgagatg ctgaagcctg
     2221 aatatttccc cgccaactac tttgccgaga gcggcgtggg ccggatcaag gaccgggtgc
     2281 tgaacagact gaacaaggcc atcaagagca acaaggccaa gaagggcgag atcatcgcct
     2341 atgacaagat gagagaagtg atggctttca tcaataactc tctgcccgtg gacgagaagc
     2401 tgaagcccaa ggattacaag agatacctgg gcatggtgag attctgggat agagaaaagg
     2461 acaatatcaa gcgcgagttc gaaacgaagg agtggagcaa gtatctgccc tccaacttct
     2521 ggaccgccaa gaacctggag agagtgtacg gactggcccg ggaaaagaac gcagagctgt
     2581 ttaacaagct gaaggccgac gtggagaaga tggacgaaag agagctggaa aagtatcaga
     2641 agatcaacga cgccaaggat ctggccaacc tgcggcggct ggccagcgac ttcggagtga
     2701 agtgggagga gaaggattgg gacgagtact ccggccagat caagaagcag atcacagatt
     2761 cccagaagct gaccatcatg aagcagagaa tcacagccgg cctgaagaag aagcacggca
     2821 tcgaaaacct gaacctgagg atcaccatcg acatcaacaa gtccagaaag gccgtgctga
     2881 atcggatcgc catccccaga ggatttgtga agcggcacat cctgggctgg caggaatccg
     2941 agaaggtgag caagaagatc agagaagccg aatgcgagat tctgctgagc aaggagtacg
     3001 aggagctgag caagcagttc tttcagagca aggactacga caagatgacc cgcatcaacg
     3061 gcctgtacga gaagaataag ctgatcgccc tgatggccgt gtatctgatg gggcagctga
     3121 gaatcctgtt caaggagcac accaagctgg acgacatcac caagaccacc gtggatttca
     3181 agatcagcga caaggtgacc gtgaagatcc ccttctccaa ctatccctcc ctggtgtaca
     3241 ccatgagcag caagtacgtg gacaatatcg gcaactacgg cttcagcaac aaggacaagg
     3301 ataagcccat tctgggcaag atcgacgtga tcgagaagca gcggatggag tttatcaagg
     3361 aggtgctggg attcgagaag tacctgtttg acgataagat catcgacaag agcaagttcg
     3421 ccgacaccgc cacccacatc agctttgccg aaatcgtgga agaactggtg gagaagggct
     3481 gggacaagga ccggctgacg aagctgaagg atgcccggaa caaggccctg cacggcgaga
     3541 tcctgaccgg caccagcttc gacgagacaa agtccctgat caacgagctg aagaagccca
     3601 agaagaagcg gaaggtgtcc tctgattata aggaccacga tggcgactac aaggatcacg
     3661 acatcgatta taaggacgat gacgataagt aaaagcttaa cttgtttatt gcagcttata
     3721 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3781 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atcactagtg
     3841 agggcctatt tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga
     3901 taattggaat taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa
     3961 agtaataatt tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata
     4021 tgcttaccgt aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac
     4081 gaaacaccga tgtcttcctg ggacgaagac aagctgtgat agacctcgat ttgtggggta
     4141 gtaacagctt tttttacgcg ttagttatta atagtaatca attacggggt cattagttca
     4201 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
     4261 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
     4321 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
     4381 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
     4441 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
     4501 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
     4561 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
     4621 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
     4681 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa
     4741 ccgtcagatc cgctagcgcc accatggtga gcaagggcga ggaggataac atggccatca
     4801 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
     4861 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     4921 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
     4981 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
     5041 ccgagggctt caagtgggag cgcgtgatga acttcgatga cggcggcgtg gtgaccgtga
     5101 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
     5161 acttcccctc cgacggcccc gtaatgcaga agaaaaccat gggctgggag gcctcctccg
     5221 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     5281 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     5341 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     5401 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     5461 agctgtacaa gtaactcttg gcccggatcc ctgtgccttc tagttgccag ccatctgttg
     5521 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     5581 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     5641 gggtggggca ggacagcaag ggggaggatt gggaaaacaa tagcaggcat gctggggatg
     5701 cggtgggctc tatgggtcga cggcgcgccc tagacgaagc agctccagcc tacacaatcg
     5761 ctcaagacgt gtaatgcttt tattatatat tagtcacgat atctataaca agaaaatata
     5821 tatataataa gttatcacgt aagtagaaca tgaaataaca atataattat cgtatgagtt
     5881 aaatcttaaa agtcacgtaa aagataatca tgcgtcattt tgactcacgc ggtcgttata
     5941 gttcaaaatc agtgacactt accgcattga caagcacgcc tcacgggagc tccaagcggc
     6001 gactgagatg tcctaaatgc acagcgacgg attcgcgcta tttagaaaga gagagcaata
     6061 tttcaagaat gcatgcgtca attttacgca gactatcttt ctagggttaa ggatccgcac
     6121 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     6181 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     6241 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     6301 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     6361 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     6421 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     6481 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     6541 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     6601 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     6661 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     6721 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     6781 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     6841 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     6901 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     6961 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     7021 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     7081 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     7141 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     7201 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     7261 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     7321 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     7381 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     7441 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     7501 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     7561 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     7621 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     7681 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     7741 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     7801 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     7861 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     7921 gttctttcct gcgt
//
