LOCUS       sequence-354636-        9137 bp    DNA     circular SYN 15-APR-2025
DEFINITION  sgRNA for CRISPR/Cas9-mediated deletion of human NRF2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9137)
  TITLE     Ristow lab plasmids
REFERENCE   2  (bases 1 to 9137)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: PX459_NRF2-exon5-2
FEATURES             Location/Qualifiers
     source          1..9137
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        443..728
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          968..1195
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1207..1216
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1216..1281
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1288..1308
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1333..5433
                     /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             1333..5433
                     /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             5434..5481
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5497..5550
                     /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             5551..6150
                     /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(5551..5570)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6047..6067
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6178..6195)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6184..6391
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6400..6540
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6400..6529
                     /label=AAV2 ITR
     rep_origin      6615..7070
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6702..6721)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6912..6933
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7087..7106)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7206..7228
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7266..7284)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7352..7456
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7457..7525,7526..8317)
                     /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(7675..7694)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8488..9076
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8977..8996
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ttgtgagatg agcctccaag gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga
      361 gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca
      421 gacaaatggc tctagaggta cccgttacat aacttacggt aaatggcccg cctggctgac
      481 cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc aatagggact ttccattgac
      541 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      601 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattgtgccc
      661 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      721 ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc
      781 cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg
      841 gggggggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg
      901 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg
      961 gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg
     1021 ccgcccgccc cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc
     1081 ttctcctccg ggctgtaatt agctgagcaa gaggtaaggg tttaagggat ggttggttgg
     1141 tggggtatta atgtttaatt acctggagca cctgcctgaa atcacttttt ttcaggttgg
     1201 accggtgcca ccatggacta taaggaccac gacggagact acaaggatca tgatattgat
     1261 tacaaagacg atgacgataa gatggcccca aagaagaagc ggaaggtcgg tatccacgga
     1321 gtcccagcag ccgacaagaa gtacagcatc ggcctggaca tcggcaccaa ctctgtgggc
     1381 tgggccgtga tcaccgacga gtacaaggtg cccagcaaga aattcaaggt gctgggcaac
     1441 accgaccggc acagcatcaa gaagaacctg atcggagccc tgctgttcga cagcggcgaa
     1501 acagccgagg ccacccggct gaagagaacc gccagaagaa gatacaccag acggaagaac
     1561 cggatctgct atctgcaaga gatcttcagc aacgagatgg ccaaggtgga cgacagcttc
     1621 ttccacagac tggaagagtc cttcctggtg gaagaggata agaagcacga gcggcacccc
     1681 atcttcggca acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac
     1741 ctgagaaaga aactggtgga cagcaccgac aaggccgacc tgcggctgat ctatctggcc
     1801 ctggcccaca tgatcaagtt ccggggccac ttcctgatcg agggcgacct gaaccccgac
     1861 aacagcgacg tggacaagct gttcatccag ctggtgcaga cctacaacca gctgttcgag
     1921 gaaaacccca tcaacgccag cggcgtggac gccaaggcca tcctgtctgc cagactgagc
     1981 aagagcagac ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg
     2041 ttcggaaacc tgattgccct gagcctgggc ctgaccccca acttcaagag caacttcgac
     2101 ctggccgagg atgccaaact gcagctgagc aaggacacct acgacgacga cctggacaac
     2161 ctgctggccc agatcggcga ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc
     2221 gacgccatcc tgctgagcga catcctgaga gtgaacaccg agatcaccaa ggcccccctg
     2281 agcgcctcta tgatcaagag atacgacgag caccaccagg acctgaccct gctgaaagct
     2341 ctcgtgcggc agcagctgcc tgagaagtac aaagagattt tcttcgacca gagcaagaac
     2401 ggctacgccg gctacattga cggcggagcc agccaggaag agttctacaa gttcatcaag
     2461 cccatcctgg aaaagatgga cggcaccgag gaactgctcg tgaagctgaa cagagaggac
     2521 ctgctgcgga agcagcggac cttcgacaac ggcagcatcc cccaccagat ccacctggga
     2581 gagctgcacg ccattctgcg gcggcaggaa gatttttacc cattcctgaa ggacaaccgg
     2641 gaaaagatcg agaagatcct gaccttccgc atcccctact acgtgggccc tctggccagg
     2701 ggaaacagca gattcgcctg gatgaccaga aagagcgagg aaaccatcac cccctggaac
     2761 ttcgaggaag tggtggacaa gggcgcttcc gcccagagct tcatcgagcg gatgaccaac
     2821 ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc acagcctgct gtacgagtac
     2881 ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga ccgagggaat gagaaagccc
     2941 gccttcctga gcggcgagca gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg
     3001 aaagtgaccg tgaagcagct gaaagaggac tacttcaaga aaatcgagtg cttcgactcc
     3061 gtggaaatct ccggcgtgga agatcggttc aacgcctccc tgggcacata ccacgatctg
     3121 ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg aaaacgagga cattctggaa
     3181 gatatcgtgc tgaccctgac actgtttgag gacagagaga tgatcgagga acggctgaaa
     3241 acctatgccc acctgttcga cgacaaagtg atgaagcagc tgaagcggcg gagatacacc
     3301 ggctggggca ggctgagccg gaagctgatc aacggcatcc gggacaagca gtccggcaag
     3361 acaatcctgg atttcctgaa gtccgacggc ttcgccaaca gaaacttcat gcagctgatc
     3421 cacgacgaca gcctgacctt taaagaggac atccagaaag cccaggtgtc cggccagggc
     3481 gatagcctgc acgagcacat tgccaatctg gccggcagcc ccgccattaa gaagggcatc
     3541 ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga tgggccggca caagcccgag
     3601 aacatcgtga tcgaaatggc cagagagaac cagaccaccc agaagggaca gaagaacagc
     3661 cgcgagagaa tgaagcggat cgaagagggc atcaaagagc tgggcagcca gatcctgaaa
     3721 gaacaccccg tggaaaacac ccagctgcag aacgagaagc tgtacctgta ctacctgcag
     3781 aatgggcggg atatgtacgt ggaccaggaa ctggacatca accggctgtc cgactacgat
     3841 gtggaccata tcgtgcctca gagctttctg aaggacgact ccatcgacaa caaggtgctg
     3901 accagaagcg acaagaaccg gggcaagagc gacaacgtgc cctccgaaga ggtcgtgaag
     3961 aagatgaaga actactggcg gcagctgctg aacgccaagc tgattaccca gagaaagttc
     4021 gacaatctga ccaaggccga gagaggcggc ctgagcgaac tggataaggc cggcttcatc
     4081 aagagacagc tggtggaaac ccggcagatc acaaagcacg tggcacagat cctggactcc
     4141 cggatgaaca ctaagtacga cgagaatgac aagctgatcc gggaagtgaa agtgatcacc
     4201 ctgaagtcca agctggtgtc cgatttccgg aaggatttcc agttttacaa agtgcgcgag
     4261 atcaacaact accaccacgc ccacgacgcc tacctgaacg ccgtcgtggg aaccgccctg
     4321 atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg gcgactacaa ggtgtacgac
     4381 gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca aggctaccgc caagtacttc
     4441 ttctacagca acatcatgaa ctttttcaag accgagatta ccctggccaa cggcgagatc
     4501 cggaagcggc ctctgatcga gacaaacggc gaaaccgggg agatcgtgtg ggataagggc
     4561 cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc aagtgaatat cgtgaaaaag
     4621 accgaggtgc agacaggcgg cttcagcaaa gagtctatcc tgcccaagag gaacagcgat
     4681 aagctgatcg ccagaaagaa ggactgggac cctaagaagt acggcggctt cgacagcccc
     4741 accgtggcct attctgtgct ggtggtggcc aaagtggaaa agggcaagtc caagaaactg
     4801 aagagtgtga aagagctgct ggggatcacc atcatggaaa gaagcagctt cgagaagaat
     4861 cccatcgact ttctggaagc caagggctac aaagaagtga aaaaggacct gatcatcaag
     4921 ctgcctaagt actccctgtt cgagctggaa aacggccgga agagaatgct ggcctctgcc
     4981 ggcgaactgc agaagggaaa cgaactggcc ctgccctcca aatatgtgaa cttcctgtac
     5041 ctggccagcc actatgagaa gctgaagggc tcccccgagg ataatgagca gaaacagctg
     5101 tttgtggaac agcacaagca ctacctggac gagatcatcg agcagatcag cgagttctcc
     5161 aagagagtga tcctggccga cgctaatctg gacaaagtgc tgtccgccta caacaagcac
     5221 cgggataagc ccatcagaga gcaggccgag aatatcatcc acctgtttac cctgaccaat
     5281 ctgggagccc ctgccgcctt caagtacttt gacaccacca tcgaccggaa gaggtacacc
     5341 agcaccaaag aggtgctgga cgccaccctg atccaccaga gcatcaccgg cctgtacgag
     5401 acacggatcg acctgtctca gctgggaggc gacaaaaggc cggcggccac gaaaaaggcc
     5461 ggccaggcaa aaaagaaaaa ggaattcggc agtggagagg gcagaggaag tctgctaaca
     5521 tgcggtgacg tcgaggagaa tcctggccca atgaccgagt acaagcccac ggtgcgcctc
     5581 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     5641 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     5701 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     5761 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     5821 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     5881 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggagtc
     5941 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     6001 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     6061 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     6121 tggtgcatga cccgcaagcc cggtgcctga gaattctaac tagagctcgc tgatcagcct
     6181 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     6241 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     6301 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     6361 attgggaaga gaatagcagg catgctgggg agcggccgca ggaaccccta gtgatggagt
     6421 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc
     6481 gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg
     6541 ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc
     6601 aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     6661 gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc
     6721 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     6781 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg
     6841 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     6901 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta
     6961 ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat
     7021 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac
     7081 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc
     7141 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     7201 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg
     7261 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     7321 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     7381 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     7441 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     7501 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     7561 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     7621 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     7681 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     7741 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     7801 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     7861 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     7921 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     7981 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     8041 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     8101 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     8161 cggtgagcgt ggaagccgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     8221 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     8281 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     8341 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     8401 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     8461 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     8521 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     8581 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     8641 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     8701 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     8761 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     8821 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     8881 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     8941 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     9001 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     9061 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     9121 gccttttgct cacatgt
//
