LOCUS       sequence-195314-        8273 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses a sgRNA and a Cas9 VQR variant that recognizes ""NGA"" PAM
            motifs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 8273)
  AUTHORS   Lambrus BG, Moyer TC, Holland AJ
  TITLE     Applying the auxin-inducible degradation system for rapid protein 
            depletion in mammalian cells.
  JOURNAL   Methods Cell Biol. 2018;144:107-135. doi: 
            10.1016/bs.mcb.2018.03.004. Epub 2018 Apr 5.
  PUBMED    29804665
REFERENCE   2  (bases 1 to 8273)
  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: "Methods 
            Cell Biol."; date: "2018"; volume: "144:107-135. doi"; pages: " 
            10.1016/bs.mcb.2018.03.004. Epub 2018 Apr 5"
COMMENT     Sequence Label: p330 VQR
FEATURES             Location/Qualifiers
     source          1..8273
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     regulatory      237..246
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             246..311
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             318..338
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(363..3761,3762..3764,3765..4361,4362..4364,4365..4367,
                     4368..4370,4371..4463)
                     /codon_start=1
                     /product="Streptococcus pyogenes Cas9 endonuclease with the
                     D1135V/R1335Q/T1337R mutations"
                     /label=Cas9 VQR
                     /note="has an altered recognition specificity for the 
                     protospacer adjacent motif (PAM) (Kleinstiver et al., 
                     2015)"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKQYRSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             4464..4511
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(4539..4556)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4545..4752
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4761..4901
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4761..4890
                     /label=AAV2 ITR
     rep_origin      4976..5431
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5063..5082)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5273..5294
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5448..5467)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5567..5589
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5627..5645)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5713..5817
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5818..5886,5887..6678)
                     /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(6036..6055)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6849..7437
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7338..7357
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        7499..7747
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7499..7519
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7670..7689
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7766..7841
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        7938..8223
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
ORIGIN
        1 gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc
       61 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
      121 ggctgtaatt agctgagcaa gaggtaaggg tttaagggat ggttggttgg tggggtatta
      181 atgtttaatt acctggagca cctgcctgaa atcacttttt ttcaggttgg accggtgcca
      241 ccatggacta taaggaccac gacggagact acaaggatca tgatattgat tacaaagacg
      301 atgacgataa gatggcccca aagaagaagc ggaaggtcgg tatccacgga gtcccagcag
      361 ccgacaagaa gtacagcatc ggcctggaca tcggcaccaa ctctgtgggc tgggccgtga
      421 tcaccgacga gtacaaggtg cccagcaaga aattcaaggt gctgggcaac accgaccggc
      481 acagcatcaa gaagaacctg atcggagccc tgctgttcga cagcggcgaa acagccgagg
      541 ccacccggct gaagagaacc gccagaagaa gatacaccag acggaagaac cggatctgct
      601 atctgcaaga gatcttcagc aacgagatgg ccaaggtgga cgacagcttc ttccacagac
      661 tggaagagtc cttcctggtg gaagaggata agaagcacga gcggcacccc atcttcggca
      721 acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac ctgagaaaga
      781 aactggtgga cagcaccgac aaggccgacc tgcggctgat ctatctggcc ctggcccaca
      841 tgatcaagtt ccggggccac ttcctgatcg agggcgacct gaaccccgac aacagcgacg
      901 tggacaagct gttcatccag ctggtgcaga cctacaacca gctgttcgag gaaaacccca
      961 tcaacgccag cggcgtggac gccaaggcca tcctgtctgc cagactgagc aagagcagac
     1021 ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg ttcggaaacc
     1081 tgattgccct gagcctgggc ctgaccccca acttcaagag caacttcgac ctggccgagg
     1141 atgccaaact gcagctgagc aaggacacct acgacgacga cctggacaac ctgctggccc
     1201 agatcggcga ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc gacgccatcc
     1261 tgctgagcga catcctgaga gtgaacaccg agatcaccaa ggcccccctg agcgcctcta
     1321 tgatcaagag atacgacgag caccaccagg acctgaccct gctgaaagct ctcgtgcggc
     1381 agcagctgcc tgagaagtac aaagagattt tcttcgacca gagcaagaac ggctacgccg
     1441 gctacattga cggcggagcc agccaggaag agttctacaa gttcatcaag cccatcctgg
     1501 aaaagatgga cggcaccgag gaactgctcg tgaagctgaa cagagaggac ctgctgcgga
     1561 agcagcggac cttcgacaac ggcagcatcc cccaccagat ccacctggga gagctgcacg
     1621 ccattctgcg gcggcaggaa gatttttacc cattcctgaa ggacaaccgg gaaaagatcg
     1681 agaagatcct gaccttccgc atcccctact acgtgggccc tctggccagg ggaaacagca
     1741 gattcgcctg gatgaccaga aagagcgagg aaaccatcac cccctggaac ttcgaggaag
     1801 tggtggacaa gggcgcttcc gcccagagct tcatcgagcg gatgaccaac ttcgataaga
     1861 acctgcccaa cgagaaggtg ctgcccaagc acagcctgct gtacgagtac ttcaccgtgt
     1921 ataacgagct gaccaaagtg aaatacgtga ccgagggaat gagaaagccc gccttcctga
     1981 gcggcgagca gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg aaagtgaccg
     2041 tgaagcagct gaaagaggac tacttcaaga aaatcgagtg cttcgactcc gtggaaatct
     2101 ccggcgtgga agatcggttc aacgcctccc tgggcacata ccacgatctg ctgaaaatta
     2161 tcaaggacaa ggacttcctg gacaatgagg aaaacgagga cattctggaa gatatagtgt
     2221 tgactcttac cctctttgaa gatcgggaaa tgattgagga aagactaaaa acatacgctc
     2281 acctgttcga cgataaggtt atgaaacagt taaagaggcg tcgctatacg ggctggggac
     2341 gattgtcgcg gaaacttatc aacgggataa gagacaagca aagtggtaaa actattctcg
     2401 attttctaaa gagcgacggc ttcgccaata ggaactttat gcagctgatc catgatgact
     2461 ctttaacctt caaagaggat atacaaaagg cacaggtttc cggacaaggg gactcattgc
     2521 acgaacatat tgcgaatctt gctggttcgc cagccatcaa aaagggcata ctccagacag
     2581 tcaaagtagt ggatgagcta gttaaggtca tgggacgtca caaaccggaa aacattgtaa
     2641 tcgagatggc acgcgaaaat caaacgactc agaaggggca aaaaaacagt cgagagcgga
     2701 tgaagagaat agaagagggt attaaagaac tgggcagcca gatcttaaag gagcatcctg
     2761 tggaaaatac ccaattgcag aacgagaaac tttacctcta ttacctacaa aatggaaggg
     2821 acatgtatgt tgatcaggaa ctggacataa accgtttatc tgattacgac gtcgatcaca
     2881 ttgtacccca atcctttttg aaggacgatt caatcgacaa taaagtgctt acacgctcgg
     2941 ataagaaccg agggaaaagt gacaatgttc caagcgagga agtcgtaaag aaaatgaaga
     3001 actattggcg gcagctccta aatgcgaaac tgataacgca aagaaagttc gataacttaa
     3061 ctaaagctga gaggggtggc ttgtctgaac ttgacaaggc cggatttatt aaacgtcagc
     3121 tcgtggaaac ccgccaaatc acaaagcatg ttgcacagat actagattcc cgaatgaata
     3181 cgaaatacga cgagaacgat aagctgattc gggaagtcaa agtaatcact ttaaagtcaa
     3241 aattggtgtc ggacttcaga aaggattttc aattctataa agttagggag ataaataact
     3301 accaccatgc gcacgacgct tatcttaatg ccgtcgtagg gaccgcactc attaagaaat
     3361 acccgaagct agaaagtgag tttgtgtatg gtgattacaa agtttatgac gtccgtaaga
     3421 tgatcgcgaa aagcgaacag gagataggca aggctacagc caaatacttc ttttattcta
     3481 acattatgaa tttctttaag acggaaatca ctctggcaaa cggagagata cgcaaacgac
     3541 ctttaattga aaccaatggg gagacaggtg aaatcgtatg ggataagggc cgggacttcg
     3601 cgacggtgag aaaagttttg tccatgcccc aagtcaacat agtaaagaaa actgaggtgc
     3661 agaccggagg gttttcaaag gaatcgattc ttccaaaaag gaatagtgat aagctcatcg
     3721 ctcgtaaaaa ggactgggac ccgaaaaagt acggtggctt cgtgagccct acagttgcct
     3781 attctgtcct agtagtggca aaagttgaga agggaaaatc caagaaactg aagtcagtca
     3841 aagaattatt ggggataacg attatggagc gctcgtcttt tgaaaagaac cccatcgact
     3901 tccttgaggc gaaaggttac aaggaagtaa aaaaggatct cataattaaa ctaccaaagt
     3961 atagtctgtt tgagttagaa aatggccgaa aacggatgtt ggctagcgcc ggagagcttc
     4021 aaaaggggaa cgaactcgca ctaccgtcta aatacgtgaa tttcctgtat ttagcgtccc
     4081 attacgagaa gttgaaaggt tcacctgaag ataacgaaca gaagcaactt tttgttgagc
     4141 agcacaaaca ttatctcgac gaaatcatag agcaaatttc ggaattcagt aagagagtca
     4201 tcctagctga tgccaatctg gacaaagtat taagcgcata caacaagcac agggataaac
     4261 ccatacgtga gcaggcggaa aatattatcc atttgtttac tcttaccaac ctcggcgctc
     4321 cagccgcatt caagtatttt gacacaacga tagatcgcaa acagtacaga tctaccaagg
     4381 aggtgctaga cgcgacactg attcaccaat ccatcacggg attatatgaa actcggatag
     4441 atttgtcaca gcttgggggt gacaaaaggc cggcggccac gaaaaaggcc ggccaggcaa
     4501 aaaagaaaaa gtaagaattc ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     4561 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     4621 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     4681 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag agaatagcag
     4741 gcatgctggg gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg
     4801 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg
     4861 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt
     4921 tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg
     4981 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     5041 cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     5101 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     5161 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg
     5221 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     5281 ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga tttataaggg
     5341 attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     5401 aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct
     5461 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     5521 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     5581 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc
     5641 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt
     5701 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     5761 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     5821 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     5881 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     5941 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     6001 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     6061 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     6121 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     6181 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     6241 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     6301 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     6361 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     6421 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     6481 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tggaagccgc
     6541 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     6601 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     6661 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     6721 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     6781 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     6841 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     6901 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6961 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     7021 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     7081 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     7141 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     7201 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     7261 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     7321 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     7381 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     7441 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgtga
     7501 gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat
     7561 aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa
     7621 gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat
     7681 gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg
     7741 aaacaccggg tcttcgagaa gacctgtttt agagctagaa atagcaagtt aaaataaggc
     7801 tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt ttagagctag
     7861 aaatagcaag ttaaaataag gctagtccgt ttttagcgcg tgcgccaatt ctgcagacaa
     7921 atggctctag aggtacccgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     7981 aacgaccccc gcccattgac gtcaatagta acgccaatag ggactttcca ttgacgtcaa
     8041 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     8101 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcattg tgcccagtac
     8161 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     8221 atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc ctc
//
