LOCUS       sequence-386827-       11344 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  gRNA targeting the AAVS1 locus in a third generation Cas9 backbone 
            with GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11344)
  AUTHORS   Girish V, Lakhani AA, Thompson SL, Scaduto CM, Brown LM, Hagenson 
            RA, Sausville EL, Mendelson BE, Kandikuppa PK, Lukow DA, Yuan ML, 
            Stevens EC, Lee SN, Schukken KM, Akalu SM, Vasudevan A, Zou C, 
            Salovska B, Li W, Smith JC, Taylor AM, Martienssen RA, Liu Y, Sun R,
            Sheltzer JM
  TITLE     Oncogene-like addiction to aneuploidy in human cancers.
  JOURNAL   Science. 2023 Jul 6:eadg4521. doi: 10.1126/science.adg4521.
  PUBMED    37410869
REFERENCE   2  (bases 1 to 11344)
  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: "Science. 
            2023 Jul 6:eadg4521. doi: 10.1126/science.adg4521."
COMMENT     Sequence Label: AAVS1_gRNA
FEATURES             Location/Qualifiers
     source          1..11344
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..231
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             232..412
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    459..584
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1077..1310
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     promoter        complement(1489..1737)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(1547..1566)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1717..1737)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    1744..1861
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1930..2141
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2161..2170
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2170..2193
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2200..2220
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2245..6345
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             2245..6345
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6346..6393
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6409..6465
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(6466..6468,6469..6471,6472..7185)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(6466..6468,6469..6471,6472..7182)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6772..6791)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7119..7140
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    7201..7789
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7254..7274)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7672..7683)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7861..8094
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8172..8293
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8209..8228)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8263..8282
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8333..8468
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8488..8507)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8489..8507)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8517..8534)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8517..8533)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8526..8548)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8675..9130
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8762..8781)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8972..8993
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9156..9260
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9261..9329,9330..10121)
                     /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(9479..9498)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10292..10880
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10781..10800
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11034..11051
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11168..11189
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11204..11209,11210..11227,11228..11234)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11242..11258
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     11247..11269
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11266..11282
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11266..11282
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11301..11321
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11303..11321
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa
       61 catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac
      121 gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa
      181 ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc
      241 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
      301 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
      361 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
      421 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
      481 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
      541 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
      601 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
      661 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
      721 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
      781 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
      841 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
      901 gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg
      961 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     1021 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     1081 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     1141 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     1201 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     1261 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     1321 ggttgctctg gaaaactcgc catttgtctc aagatctaga attcaaaaaa agcaccgact
     1381 cggtgccact ttgctgtttc cagcaaagtt gataacggac tagccttatt taaacttgct
     1441 atgctgtttc cagcatagct cttaaacacg ccgacggtat cagcgcccgg tgtttcgtcc
     1501 tttccacaag atatataaag ccaagaaatc gaaatacttt caagttacgg taagcatatg
     1561 atagtccatt ttaaaacata attttaaaac tgcaaactac ccaagaaatt attactttct
     1621 acgtcacgta ttttgtacta atatctttgt gtttacagtc aaattaattc taattatctc
     1681 tctaacagcc ttgtatcgta tatgcaaata tgaaggaatc atgggaaata ggccctccca
     1741 caattttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1801 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     1861 tcgggtttat tacagggaca gcagagatcc actttggcgc cggctcgagt ggctccggtg
     1921 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
     1981 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
     2041 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
     2101 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtgtcgtga cgcgggatcc
     2161 gccaccatgg attacaaaga cgatgacgat aagatggccc caaagaagaa gcggaaggtc
     2221 ggtatccacg gagtcccagc agccgacaag aagtacagca tcggcctgga catcggcacc
     2281 aactctgtgg gctgggccgt gatcaccgac gagtacaagg tgcccagcaa gaaattcaag
     2341 gtgctgggca acaccgaccg gcacagcatc aagaagaacc tgatcggagc cctgctgttc
     2401 gacagcggcg aaacagccga ggccacccgg ctgaagagaa ccgccagaag aagatacacc
     2461 agacggaaga accggatctg ctatctgcaa gagatcttca gcaacgagat ggccaaggtg
     2521 gacgacagct tcttccacag actggaagag tccttcctgg tggaagagga taagaagcac
     2581 gagcggcacc ccatcttcgg caacatcgtg gacgaggtgg cctaccacga gaagtacccc
     2641 accatctacc acctgagaaa gaaactggtg gacagcaccg acaaggccga cctgcggctg
     2701 atctatctgg ccctggccca catgatcaag ttccggggcc acttcctgat cgagggcgac
     2761 ctgaaccccg acaacagcga cgtggacaag ctgttcatcc agctggtgca gacctacaac
     2821 cagctgttcg aggaaaaccc catcaacgcc agcggcgtgg acgccaaggc catcctgtct
     2881 gccagactga gcaagagcag acggctggaa aatctgatcg cccagctgcc cggcgagaag
     2941 aagaatggcc tgttcggaaa cctgattgcc ctgagcctgg gcctgacccc caacttcaag
     3001 agcaacttcg acctggccga ggatgccaaa ctgcagctga gcaaggacac ctacgacgac
     3061 gacctggaca acctgctggc ccagatcggc gaccagtacg ccgacctgtt tctggccgcc
     3121 aagaacctgt ccgacgccat cctgctgagc gacatcctga gagtgaacac cgagatcacc
     3181 aaggcccccc tgagcgcctc tatgatcaag agatacgacg agcaccacca ggacctgacc
     3241 ctgctgaaag ctctcgtgcg gcagcagctg cctgagaagt acaaagagat tttcttcgac
     3301 cagagcaaga acggctacgc cggctacatt gacggcggag ccagccagga agagttctac
     3361 aagttcatca agcccatcct ggaaaagatg gacggcaccg aggaactgct cgtgaagctg
     3421 aacagagagg acctgctgcg gaagcagcgg accttcgaca acggcagcat cccccaccag
     3481 atccacctgg gagagctgca cgccattctg cggcggcagg aagattttta cccattcctg
     3541 aaggacaacc gggaaaagat cgagaagatc ctgaccttcc gcatccccta ctacgtgggc
     3601 cctctggcca ggggaaacag cagattcgcc tggatgacca gaaagagcga ggaaaccatc
     3661 accccctgga acttcgagga agtggtggac aagggcgctt ccgcccagag cttcatcgag
     3721 cggatgacca acttcgataa gaacctgccc aacgagaagg tgctgcccaa gcacagcctg
     3781 ctgtacgagt acttcaccgt gtataacgag ctgaccaaag tgaaatacgt gaccgaggga
     3841 atgagaaagc ccgccttcct gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc
     3901 aagaccaacc ggaaagtgac cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag
     3961 tgcttcgact ccgtggaaat ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca
     4021 taccacgatc tgctgaaaat tatcaaggac aaggacttcc tggacaatga ggaaaacgag
     4081 gacattctgg aagatatcgt gctgaccctg acactgtttg aggacagaga gatgatcgag
     4141 gaacggctga aaacctatgc ccacctgttc gacgacaaag tgatgaagca gctgaagcgg
     4201 cggagataca ccggctgggg caggctgagc cggaagctga tcaacggcat ccgggacaag
     4261 cagtccggca agacaatcct ggatttcctg aagtccgacg gcttcgccaa cagaaacttc
     4321 atgcagctga tccacgacga cagcctgacc tttaaagagg acatccagaa agcccaggtg
     4381 tccggccagg gcgatagcct gcacgagcac attgccaatc tggccggcag ccccgccatt
     4441 aagaagggca tcctgcagac agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg
     4501 cacaagcccg agaacatcgt gatcgaaatg gccagagaga accagaccac ccagaaggga
     4561 cagaagaaca gccgcgagag aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc
     4621 cagatcctga aagaacaccc cgtggaaaac acccagctgc agaacgagaa gctgtacctg
     4681 tactacctgc agaatgggcg ggatatgtac gtggaccagg aactggacat caaccggctg
     4741 tccgactacg atgtggacca tatcgtgcct cagagctttc tgaaggacga ctccatcgac
     4801 aacaaggtgc tgaccagaag cgacaagaac cggggcaaga gcgacaacgt gccctccgaa
     4861 gaggtcgtga agaagatgaa gaactactgg cggcagctgc tgaacgccaa gctgattacc
     4921 cagagaaagt tcgacaatct gaccaaggcc gagagaggcg gcctgagcga actggataag
     4981 gccggcttca tcaagagaca gctggtggaa acccggcaga tcacaaagca cgtggcacag
     5041 atcctggact cccggatgaa cactaagtac gacgagaatg acaagctgat ccgggaagtg
     5101 aaagtgatca ccctgaagtc caagctggtg tccgatttcc ggaaggattt ccagttttac
     5161 aaagtgcgcg agatcaacaa ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg
     5221 ggaaccgccc tgatcaaaaa gtaccctaag ctggaaagcg agttcgtgta cggcgactac
     5281 aaggtgtacg acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc
     5341 gccaagtact tcttctacag caacatcatg aactttttca agaccgagat taccctggcc
     5401 aacggcgaga tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg
     5461 tgggataagg gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat
     5521 atcgtgaaaa agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag
     5581 aggaacagcg ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc
     5641 ttcgacagcc ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag
     5701 tccaagaaac tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc
     5761 ttcgagaaga atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac
     5821 ctgatcatca agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg
     5881 ctggcctctg ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg
     5941 aacttcctgt acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag
     6001 cagaaacagc tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc
     6061 agcgagttct ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc
     6121 tacaacaagc accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt
     6181 accctgacca atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg
     6241 aagaggtaca ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc
     6301 ggcctgtacg agacacggat cgacctgtct cagctgggag gcgacaagcg tcctgctgct
     6361 actaagaaag ctggtcaagc taagaaaaag aaagctagcg gcagcggcgc caccaacttc
     6421 agcctgctga agcaggccgg cgacgtggag gagaaccccg gccccatggt gagcaagggc
     6481 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     6541 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     6601 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     6661 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     6721 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     6781 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     6841 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     6901 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     6961 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     7021 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     7081 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     7141 accgccgccg ggatcactct cggcatggac gagctgtaca agtagacgcg ttaagtcgac
     7201 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     7261 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     7321 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     7381 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     7441 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     7501 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     7561 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     7621 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     7681 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     7741 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcg tcgactttaa
     7801 gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac
     7861 tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc
     7921 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
     7981 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
     8041 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtacgt
     8101 atagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag
     8161 agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     8221 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     8281 tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc ccatcccgcc
     8341 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     8401 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     8461 tggaggccta gggacgtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg
     8521 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     8581 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     8641 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     8701 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     8761 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     8821 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     8881 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     8941 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     9001 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     9061 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     9121 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     9181 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     9241 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     9301 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     9361 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     9421 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     9481 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     9541 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     9601 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     9661 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     9721 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     9781 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     9841 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     9901 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     9961 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
    10021 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
    10081 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
    10141 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
    10201 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
    10261 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
    10321 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
    10381 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
    10441 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
    10501 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
    10561 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
    10621 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
    10681 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
    10741 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
    10801 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
    10861 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
    10921 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
    10981 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
    11041 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
    11101 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
    11161 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
    11221 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
    11281 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctgc
    11341 aagc
//
