LOCUS       sequence-232547-        6890 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway Donor; RBFOX1N-dCasRx-C ORF; No promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6890)
  AUTHORS   Du M, Jillette N, Zhu JJ, Li S, Cheng AW
  TITLE     CRISPR artificial splicing factors.
  JOURNAL   Nat Commun. 2020 Jun 12;11(1):2973. doi: 10.1038/s41467-020-16806-4.
  PUBMED    32532987
REFERENCE   2  (bases 1 to 6890)
  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: "Nat 
            Commun."; date: "2020-06-12"; volume: "11(1):2973. doi"; pages: " 
            10.1038/s41467-020-16806-4"
COMMENT     Sequence Label: pAC1823_pCR8-RBFOX1N-dCasRx-C
FEATURES             Location/Qualifiers
     source          1..6890
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(157..184)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(276..362)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     411..433
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     425..442
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     426..442
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    458..557
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             959..979
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             989..3886
                     /codon_start=1
                     /product="RNA-targeting Type VI CRISPR protein from 
                     Ruminococcus flavefaciens strain XPD3002 (Konermann et al.,
                     2018)"
                     /label=RfxCas13d
                     /translation="IEKKKSFAKGMGVKSTLVSGSKVYMTTFAEGSDARLEKIVEGDSI
                     RSVNEGEAFSAEMADKNAGYKIGNAKFSHPKGYAVVANNPLYTGPVQQDMLGLKETLEK
                     RYFGESADGNDNICIQVIHNILDIEKILAEYITNAAYAVNNISGLDKDIIGFGKFSTVY
                     TYDEFKDPEHHRAAFNNNDKLINAIKAQYDEFDNFLDNPRLGYFGQAFFSKEGRNYIIN
                     YGNECYDILALLSGLAHWVVANNEEESRISRTWLYNLDKNLDNEYISTLNYLYDRITNE
                     LTNSFSKNSAANVNYIAETLGINPAEFAEQYFRFSIMKEQKNLGFNITKLREVMLDRKD
                     MSEIRKNHKVFDSIRTKVYTMMDFVIYRYYIEEDAKVAAANKSLPDNEKSLSEKDIFVI
                     NLRGSFNDDQKDALYYDEANRIWRKLENIMHNIKEFRGNKTREYKKKDAPRLPRILPAG
                     RDVSAFSKLMYALTMFLDGKEINDLLTTLINKFDNIQSFLKVMPLIGVNAKFVEEYAFF
                     KDSAKIADELRLIKSFARMGEPIADARRAMYIDAIRILGTNLSYDELKALADTFSLDEN
                     GNKLKKGKHGMRNFIINNVISNKRFHYLIRYGDPAHLHEIAKNEAVVKFVLGRIADIQK
                     KQGQNGKNQIDRYYETCIGKDKGKSVSEKVDALTKIITGMNYDQFDKKRSVIEDTGREN
                     AEREKFKKIISLYLTVIYHILKNIVNINARYVIGFHCVERDAQLYKEKGYDINLKKLEE
                     KGFSSVTKLCAGIDETAPDKRKDVEKEMAERAKESIDSLESANPKLYANYIKYSDEKKA
                     EEFTRQINREKAKTALNAYLRNTKWNVIIREDLLRIDNKTCTLFANKAVALEVARYVHA
                     YINDIAEVNSYFQLYHYIMQRIIMNERYEKSSGKVSEYFDAVNDEKKYNDRLLKLLCVP
                     FGYCIPRFKNLSIEALFDRNEAAKFDKEKKKVSGNS"
     CDS             3896..3916
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3926..3952
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    complement(4658..4757)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4774..4793)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4775..4793)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4798..4814)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4798..4814)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5245..6036
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      6129..6717
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6618..6637
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6871..6888
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
       61 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatac
      121 gcgtaccgct agccaggaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc
      181 ttctgcttag tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc
      241 cgttgcttca caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc
      301 accgacaaac aacagataaa acgaaaggcc cagtcttccg actgagcctt tcgttttatt
      361 tgatgcctgg cagttcccta ctctcgcgtt aacgctagca tggatgtttt cccagtcacg
      421 acgttgtaaa acgacggcca gtcttaagct cgggccccaa ataatgattt tattttgact
      481 gatagtgacc tgttcgttgc aacaaattga tgagcaatgc ttttttataa tgccaacttt
      541 gtacaaaaaa gcaggctccg aattcaccgg tgccgccacc atgaattgcg agcgggaaca
      601 gctcaggggc aatcaagaag cggcagctgc acctgacaca atggcacagc cgtatgcgag
      661 tgctcaattt gcgccacccc agaatggcat acccgcagaa tacacagcgc cgcatcctca
      721 ccccgcccct gaatacactg ggcaaactac cgttcctgaa cacacgctta acctctatcc
      781 tcctgctcag actcactcag aacagagtcc agcagataca agtgcgcaga cagtctctgg
      841 cactgcgacc cagaccgatg acgcagctcc tactgatggc cagccacaaa ctcaaccctc
      901 tgaaaacact gaaaataaaa gccaacccaa gggcggtgga ggctccgggc gcgccagccc
      961 caagaagaag agaaaggtgg aggccagcat cgaaaaaaaa aagtccttcg ccaagggcat
     1021 gggcgtgaag tccacactcg tgtccggctc caaagtgtac atgacaacct tcgccgaagg
     1081 cagcgacgcc aggctggaaa agatcgtgga gggcgacagc atcaggagcg tgaatgaggg
     1141 cgaggccttc agcgctgaaa tggccgataa aaacgccggc tataagatcg gcaacgccaa
     1201 attcagccat cctaagggct acgccgtggt ggctaacaac cctctgtata caggacccgt
     1261 ccagcaggat atgctcggcc tgaaggaaac tctggaaaag aggtacttcg gcgagagcgc
     1321 tgatggcaat gacaatattt gtatccaggt gatccataac atcctggaca ttgaaaaaat
     1381 cctcgccgaa tacattacca acgccgccta cgccgtcaac aatatctccg gcctggataa
     1441 ggacattatt ggattcggca agttctccac agtgtatacc tacgacgaat tcaaagaccc
     1501 cgagcaccat agggccgctt tcaacaataa cgataagctc atcaacgcca tcaaggccca
     1561 gtatgacgag ttcgacaact tcctcgataa ccccagactc ggctatttcg gccaggcctt
     1621 tttcagcaag gagggcagaa attacatcat caattacggc aacgaatgct atgacattct
     1681 ggccctcctg agcggactgg cgcactgggt ggtcgctaac aacgaagaag agtccaggat
     1741 ctccaggacc tggctctaca acctcgataa gaacctcgac aacgaataca tctccaccct
     1801 caactacctc tacgacagga tcaccaatga gctgaccaac tccttctcca agaactccgc
     1861 cgccaacgtg aactatattg ccgaaactct gggaatcaac cctgccgaat tcgccgaaca
     1921 atatttcaga ttcagcatta tgaaagagca gaaaaacctc ggattcaata tcaccaagct
     1981 cagggaagtg atgctggaca ggaaggatat gtccgagatc aggaaaaatc ataaggtgtt
     2041 cgactccatc aggaccaagg tctacaccat gatggacttt gtgatttata ggtattacat
     2101 cgaagaggat gccaaggtgg ctgccgccaa taagtccctc cccgataatg agaagtccct
     2161 gagcgagaag gatatctttg tgattaacct gaggggctcc ttcaacgacg accagaagga
     2221 tgccctctac tacgatgaag ctaatagaat ttggagaaag ctcgaaaata tcatgcacaa
     2281 catcaaggaa tttaggggaa acaagacaag agagtataag aagaaggacg cccctagact
     2341 gcccagaatc ctgcccgctg gccgtgatgt ttccgccttc agcaaactca tgtatgccct
     2401 gaccatgttc ctggatggca aggagatcaa cgacctcctg accaccctga ttaataaatt
     2461 cgataacatc cagagcttcc tgaaggtgat gcctctcatc ggagtcaacg ctaagttcgt
     2521 ggaggaatac gcctttttca aagactccgc caagatcgcc gatgagctga ggctgatcaa
     2581 gtccttcgct agaatgggag aacctattgc cgatgccagg agggccatgt atatcgacgc
     2641 catccgtatt ttaggaacca acctgtccta tgatgagctc aaggccctcg ccgacacctt
     2701 ttccctggac gagaacggaa acaagctcaa gaaaggcaag cacggcatga gaaatttcat
     2761 tattaataac gtgatcagca ataaaaggtt ccactacctg atcagatacg gtgatcctgc
     2821 ccacctccat gagatcgcca aaaacgaggc cgtggtgaag ttcgtgctcg gcaggatcgc
     2881 tgacatccag aaaaaacagg gccagaacgg caagaaccag atcgacaggt actacgaaac
     2941 ttgtatcgga aaggataagg gcaagagcgt gagcgaaaag gtggacgctc tcacaaagat
     3001 catcaccgga atgaactacg accaattcga caagaaaagg agcgtcattg aggacaccgg
     3061 cagggaaaac gccgagaggg agaagtttaa aaagatcatc agcctgtacc tcaccgtgat
     3121 ctaccacatc ctcaagaata ttgtcaatat caacgccagg tacgtcatcg gattccattg
     3181 cgtcgagcgt gatgctcaac tgtacaagga gaaaggctac gacatcaatc tcaagaaact
     3241 ggaagagaag ggattcagct ccgtcaccaa gctctgcgct ggcattgatg aaactgcccc
     3301 cgataagaga aaggacgtgg aaaaggagat ggctgaaaga gccaaggaga gcattgacag
     3361 cctcgagagc gccaacccca agctgtatgc caattacatc aaatacagcg acgagaagaa
     3421 agccgaggag ttcaccaggc agattaacag ggagaaggcc aaaaccgccc tgaacgccta
     3481 cctgaggaac accaagtgga atgtgatcat cagggaggac ctcctgagaa ttgacaacaa
     3541 gacatgtacc ctgttcgcaa acaaggccgt cgccctggaa gtggccaggt atgtccacgc
     3601 ctatatcaac gacattgccg aggtcaattc ctacttccaa ctgtaccatt acatcatgca
     3661 gagaattatc atgaatgaga ggtacgagaa aagcagcgga aaggtgtccg agtacttcga
     3721 cgctgtgaat gacgagaaga agtacaacga taggctcctg aaactgctgt gtgtgccttt
     3781 cggctactgt atccccaggt ttaagaacct gagcatcgag gccctgttcg ataggaacga
     3841 ggccgccaag ttcgacaagg agaaaaagaa ggtgtccggc aattccggat ccggacctaa
     3901 gaaaaagagg aaggtggcgg ccgcttaccc atacgatgtt ccagattacg ctggcggccg
     3961 cgggggaggc ggttccggtg gcggcggaag cggaggtgga ggatcagggc cggccaatgc
     4021 cacggctagg gtcatgacaa acaagaagac ggttaatccg tataccaacg ggtggaaact
     4081 taatcccgtt gtcggcgctg tctattctcc cgagttctat gctggtacag tcttgctgtg
     4141 ccaagccaat caagaggggt cttcaatgta cagtgctccg agcagcttgg tttacacaag
     4201 cgcgatgcca ggatttcctt accctgcggc aacggccgcg gctgcgtatc ggggtgccca
     4261 ccttcgcggg cggggccgaa cagtttacaa cacgttcagg gccgctgctc cacctccccc
     4321 gattcccgcg tacggaggtg ttgtctacca agacgggttt tatggagccg acatctacgg
     4381 tggttacgcc gcatatcggt atgcacaacc aacccccgcg accgcggcag cgtacagcga
     4441 ctcctacgga cgcgtgtacg ctgctgatcc ttaccaccac gccctggccc ccgccccaac
     4501 gtatggtgtc ggtgctatga atgcgttcgc gcccctgacc gatgcaaaaa ccaggtcaca
     4561 tgccgacgac gttggattgg tattgagctc tttgcaagca tccatttacc gggggggata
     4621 caaccgcttc gcgccttatt gattaattaa gaattcgacc cagctttctt gtacaaagtt
     4681 ggcattataa aaaataattg ctcatcaatt tgttgcaacg aacaggtcac tatcagtcaa
     4741 aataaaatca ttatttgcca tccagctgat atcccctata gtgagtcgta ttacatggtc
     4801 atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa
     4861 gataaaaata tatcatcatg cctcctctag accagccagg acagaaatgc ctcgacttcg
     4921 ctgctaccca aggttgccgg gtgacgcaca ccgtggaaac ggatgaaggc acgaacccag
     4981 tggacataag cctgttcggt tcgtaagctg taatgcaagt agcgtatgcg ctcacgcaac
     5041 tggtccagaa ccttgaccga acgcagcggt ggtaacggcg cagtggcggt tttcatggct
     5101 tgttatgact gtttttttgg ggtacagtct atgcctcggg catccaagca gcaagcgcgt
     5161 tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg atgttacgca gcagggcagt
     5221 cgccctaaaa caaagttaaa cattatgagg gaagcggtga tcgccgaagt atcgactcaa
     5281 ctatcagagg tagttggcgt catcgagcgc catctcgaac cgacgttgct ggccgtacat
     5341 ttgtacggct ccgcagtgga tggcggcctg aagccacaca gtgatattga tttgctggtt
     5401 acggtgaccg taaggcttga tgaaacaacg cggcgagctt tgatcaacga ccttttggaa
     5461 acttcggctt cccctggaga gagcgagatt ctccgcgctg tagaagtcac cattgttgtg
     5521 cacgacgaca tcattccgtg gcgttatcca gctaagcgcg aactgcaatt tggagaatgg
     5581 cagcgcaatg acattcttgc aggtatcttc gagccagcca cgatcgacat tgatctggct
     5641 atcttgctga caaaagcaag agaacatagc gttgccttgg taggtccagc ggcggaggaa
     5701 ctctttgatc cggttcctga acaggatcta tttgaggcgc taaatgaaac cttaacgcta
     5761 tggaactcgc cgcccgactg ggctggcgat gagcgaaatg tagtgcttac gttgtcccgc
     5821 atttggtaca gcgcagtaac cggcaaaatc gcgccgaagg atgtcgctgc cgactgggca
     5881 atggagcgcc tgccggccca gtatcagccc gtcatacttg aagctagaca ggcttatctt
     5941 ggacaagaag aagatcgctt ggcctcgcgc gcagatcagt tggaagaatt tgtccactac
     6001 gtgaaaggcg agatcaccaa ggtagtcggc aaataaccct cgagccaccc atgaccaaaa
     6061 tcccttaacg tgagttacgc gtcgttccac tgagcgtcag accccgtaga aaagatcaaa
     6121 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     6181 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6241 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     6301 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     6361 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     6421 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     6481 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     6541 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     6601 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     6661 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     6721 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     6781 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     6841 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
//
