LOCUS       sequence-230651-        9175 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR plasmid for expression of Cas9 nicakse (D10A) and gRNA 
            targeting human transcription factor DEAF1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9175)
  TITLE     White lab ENCODE plasmids to tag human transcription factors with 
            GFP
REFERENCE   2  (bases 1 to 9175)
  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"
COMMENT     Sequence Label: pDEAF1.2.2-gDNA
FEATURES             Location/Qualifiers
     source          1..9175
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        168..453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        455..730
                     /label=chicken beta-actin promoter
     intron          731..958
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      970..979
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             979..1044
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1051..1071
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1096..1119,1120..1122,1123..5196)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1096..5196
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5197..5244
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5260..5313
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             5314..5913
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5314..5333)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5810..5830
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(5941..5958)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5947..6154
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6163..6303
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6163..6292
                     /label=AAV2 ITR
     rep_origin      6378..6833
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6465..6484)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6675..6696
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6850..6869)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6969..6991
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7029..7047)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7115..7219
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7220..7288,7289..8080)
                     /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(7438..7457)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8251..8839
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8740..8759
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        8901..9149
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     8901..8921
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     9072..9091
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        join(9171..9175,1..71)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
       61 cgagtcggtg cttttttgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
      121 ttttagcgcg tgcgccaatt ctgcagacaa atggctctag aggtacccgt tacataactt
      181 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaatagta
      241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      361 aaatggcccg cctggcattg tgcccagtac atgaccttat gggactttcc tacttggcag
      421 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
      481 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
      541 ttttgtgcag cgatgggggc gggggggggg ggggcgcgcg ccmggcgggg cggggcgggg
      601 cgaggggcgg ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct
      661 ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc
      721 gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc
      781 gcgccgcccg ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg
      841 cccttctcct ccgggctgta attagctgag caagaggtaa gggtttaagg gatggttggt
      901 tggtggggta ttaatgttta attacctgga gcacctgcct gaaatcactt tttttcaggt
      961 tggaccggtg ccaccatgga ctataaggac cacgacggag actacaagga tcatgatatt
     1021 gattacaaag acgatgacga taagatggcc ccaaagaaga agcggaaggt cggtatccac
     1081 ggagtcccag cagccgacaa gaagtacagc atcggcctgg ccatcggcac caactctgtg
     1141 ggctgggccg tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc
     1201 aacaccgacc ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc
     1261 gaaacagccg aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag
     1321 aaccggatct gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc
     1381 ttcttccaca gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac
     1441 cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac
     1501 cacctgagaa agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg
     1561 gccctggccc acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc
     1621 gacaacagcg acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc
     1681 gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg
     1741 agcaagagca gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc
     1801 ctgttcggca acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc
     1861 gacctggccg aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac
     1921 aacctgctgg cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg
     1981 tccgacgcca tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc
     2041 ctgagcgcct ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa
     2101 gctctcgtgc ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag
     2161 aacggctacg ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc
     2221 aagcccatcc tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag
     2281 gacctgctgc ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg
     2341 ggagagctgc acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac
     2401 cgggaaaaga tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc
     2461 aggggaaaca gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg
     2521 aacttcgagg aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc
     2581 aacttcgata agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag
     2641 tacttcaccg tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag
     2701 cccgccttcc tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac
     2761 cggaaagtga ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac
     2821 tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat
     2881 ctgctgaaaa ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg
     2941 gaagatatcg tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg
     3001 aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac
     3061 accggctggg gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc
     3121 aagacaatcc tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg
     3181 atccacgacg acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag
     3241 ggcgatagcc tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc
     3301 atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc
     3361 gagaacatcg tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac
     3421 agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg
     3481 aaagaacacc ccgtggaaaa cacccagctg cagaacgaga agctgtacct gtactacctg
     3541 cagaatgggc gggatatgta cgtggaccag gaactggaca tcaaccggct gtccgactac
     3601 gatgtggacc atatcgtgcc tcagagcttt ctgaaggacg actccatcga caacaaggtg
     3661 ctgaccagaa gcgacaagaa ccggggcaag agcgacaacg tgccctccga agaggtcgtg
     3721 aagaagatga agaactactg gcggcagctg ctgaacgcca agctgattac ccagagaaag
     3781 ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg aactggataa ggccggcttc
     3841 atcaagagac agctggtgga aacccggcag atcacaaagc acgtggcaca gatcctggac
     3901 tcccggatga acactaagta cgacgagaat gacaagctga tccgggaagt gaaagtgatc
     3961 accctgaagt ccaagctggt gtccgatttc cggaaggatt tccagtttta caaagtgcgc
     4021 gagatcaaca actaccacca cgcccacgac gcctacctga acgccgtcgt gggaaccgcc
     4081 ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt acggcgacta caaggtgtac
     4141 gacgtgcgga agatgatcgc caagagcgag caggaaatcg gcaaggctac cgccaagtac
     4201 ttcttctaca gcaacatcat gaactttttc aagaccgaga ttaccctggc caacggcgag
     4261 atccggaagc ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt gtgggataag
     4321 ggccgggatt ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa tatcgtgaaa
     4381 aagaccgagg tgcagacagg cggcttcagc aaagagtcta tcctgcccaa gaggaacagc
     4441 gataagctga tcgccagaaa gaaggactgg gaccctaaga agtacggcgg cttcgacagc
     4501 cccaccgtgg cctattctgt gctggtggtg gccaaagtgg aaaagggcaa gtccaagaaa
     4561 ctgaagagtg tgaaagagct gctggggatc accatcatgg aaagaagcag cttcgagaag
     4621 aatcccatcg actttctgga agccaagggc tacaaagaag tgaaaaagga cctgatcatc
     4681 aagctgccta agtactccct gttcgagctg gaaaacggcc ggaagagaat gctggcctct
     4741 gccggcgaac tgcagaaggg aaacgaactg gccctgccct ccaaatatgt gaacttcctg
     4801 tacctggcca gccactatga gaagctgaag ggctcccccg aggataatga gcagaaacag
     4861 ctgtttgtgg aacagcacaa gcactacctg gacgagatca tcgagcagat cagcgagttc
     4921 tccaagagag tgatcctggc cgacgctaat ctggacaaag tgctgtccgc ctacaacaag
     4981 caccgggata agcccatcag agagcaggcc gagaatatca tccacctgtt taccctgacc
     5041 aatctgggag cccctgccgc cttcaagtac tttgacacca ccatcgaccg gaagaggtac
     5101 accagcacca aagaggtgct ggacgccacc ctgatccacc agagcatcac cggcctgtac
     5161 gagacacgga tcgacctgtc tcagctggga ggcgacaaaa ggccggcggc cacgaaaaag
     5221 gccggccagg caaaaaagaa aaaggaattc ggcagtggag agggcagagg aagtctgcta
     5281 acatgcggtg acgtcgagga gaatcctggc ccaatgaccg agtacaagcc cacggtgcgc
     5341 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     5401 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     5461 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     5521 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     5581 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     5641 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga
     5701 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     5761 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     5821 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     5881 acctggtgca tgacccgcaa gcccggtgcc tgagaattct aactagagct cgctgatcag
     5941 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     6001 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     6061 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     6121 aggattggga agagaatagc aggcatgctg gggagcggcc gcaggaaccc ctagtgatgg
     6181 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     6241 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc
     6301 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     6361 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     6421 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     6481 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     6541 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     6601 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     6661 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     6721 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     6781 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     6841 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     6901 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     6961 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     7021 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     7081 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     7141 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     7201 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     7261 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     7321 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     7381 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     7441 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     7501 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     7561 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     7621 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     7681 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     7741 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     7801 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     7861 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     7921 agccggtgag cgtggaagcc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     7981 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     8041 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     8101 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     8161 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     8221 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     8281 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     8341 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     8401 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     8461 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     8521 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     8581 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     8641 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     8701 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     8761 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     8821 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     8881 ctggcctttt gctcacatgt gagggcctat ttcccatgat tccttcatat ttgcatatac
     8941 gatacaaggc tgttagagag ataattggaa ttaatttgac tgtaaacaca aagatattag
     9001 tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt ttaaaattat
     9061 gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat ttcttggctt
     9121 tatatatctt gtggaaagga cgaaacaccg gaggctccat cggccgccct gtttt
//
