LOCUS       sequence-237582-        8446 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses sgRNA targeting mouse Eif4a1 and Cas9 nickase in mammalian
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8446)
  AUTHORS   Ochiai H, Hayashi T, Umeda M, Yoshimura M, Harada A, Shimizu Y, 
            Nakano K, Saitoh N, Liu Z, Yamamoto T, Okamura T, Ohkawa Y, Kimura 
            H, Nikaido I
  TITLE     Genome-wide kinetic properties of transcriptional bursting in mouse 
            embryonic stem cells.
  JOURNAL   Sci Adv. 2020 Jun 17;6(25):eaaz6699. doi: 10.1126/sciadv.aaz6699. 
            eCollection 2020 Jun.
  PUBMED    32596448
REFERENCE   2  (bases 1 to 8446)
  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: "Sci Adv."; 
            date: "2020-06-17"; volume: "6(25):eaaz6699. doi"; pages: " 
            10.1126/sciadv.aaz6699. eCollection 2020 Jun"
COMMENT     Sequence Label: px335-sgRNA-EF(Cas9n)-Eif4a1-R
FEATURES             Location/Qualifiers
     source          1..8446
                     /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"
     enhancer        453..738
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        740..1017
                     /label=chicken beta-actin promoter
     intron          1018..1245
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1266..1292
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1296..1316
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1326..1349,1350..1352,1353..5426)
                     /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             1326..5426
                     /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             5430..5450
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5487..5504)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5493..5700
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5709..5849
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5709..5838
                     /label=AAV2 ITR
     rep_origin      5924..6379
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6011..6030)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6221..6242
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6396..6415)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6515..6537
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6575..6593)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6661..6765
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6766..6834,6835..7626)
                     /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(6984..7003)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7797..8385
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8286..8305
                     /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 gtcaggattc tcggtaagaa gtttaagagc tatgctggaa acagcatagc
      301 aagtttaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt
      361 ttgttttaga gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc
      421 caattctgca gacaaatggc tctagaggta cccgttacat aacttacggt aaatggcccg
      481 cctggctgac cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc aatagggact
      541 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      601 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      661 cattgtgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      721 gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc
      781 cccccctccc cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg
      841 ggggcggggg gggggggggg gcgcgcgccr ggcggggcgg ggsggggggg gggggggggg
      901 ggggcgrggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
      961 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
     1021 gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc
     1081 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
     1141 ggctgtaatt agctgagcaa gaggtaaggg tttaagggat ggttggttgg tggggtatta
     1201 atgtttaatt acctggagca cctgcctgaa atcacttttt ttcaggttgg accggtgcca
     1261 ccatgtaccc atacgatgtt ccagattacg cttcgccgaa gaaaaagcgc aaggtcgaag
     1321 cgtccgacaa gaagtacagc atcggcctgg ccatcggcac caactctgtg ggctgggccg
     1381 tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc aacaccgacc
     1441 ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc gaaacagccg
     1501 aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag aaccggatct
     1561 gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc ttcttccaca
     1621 gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac cccatcttcg
     1681 gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac cacctgagaa
     1741 agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg gccctggccc
     1801 acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc gacaacagcg
     1861 acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc gaggaaaacc
     1921 ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg agcaagagca
     1981 gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc ctgttcggca
     2041 acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc gacctggccg
     2101 aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac aacctgctgg
     2161 cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg tccgacgcca
     2221 tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc ctgagcgcct
     2281 ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa gctctcgtgc
     2341 ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag aacggctacg
     2401 ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc aagcccatcc
     2461 tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag gacctgctgc
     2521 ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg ggagagctgc
     2581 acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac cgggaaaaga
     2641 tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc aggggaaaca
     2701 gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg aacttcgagg
     2761 aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc aacttcgata
     2821 agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag tacttcaccg
     2881 tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc
     2941 tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac cggaaagtga
     3001 ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac tccgtggaaa
     3061 tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa
     3121 ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg gaagatatcg
     3181 tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg aaaacctatg
     3241 cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac accggctggg
     3301 gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc aagacaatcc
     3361 tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg atccacgacg
     3421 acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag ggcgatagcc
     3481 tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc atcctgcaga
     3541 cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg
     3601 tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac agccgcgaga
     3661 gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg aaagaacacc
     3721 ccgtggaaaa cacccagctg cagaacgaga agctgtacct gtactacctg cagaatgggc
     3781 gggatatgta cgtggaccag gaactggaca tcaaccggct gtccgactac gatgtggacc
     3841 atatcgtgcc tcagagcttt ctgaaggacg actccatcga caacaaggtg ctgaccagaa
     3901 gcgacaagaa ccggggcaag agcgacaacg tgccctccga agaggtcgtg aagaagatga
     3961 agaactactg gcggcagctg ctgaacgcca agctgattac ccagagaaag ttcgacaatc
     4021 tgaccaaggc cgagagaggc ggcctgagcg aactggataa ggccggcttc atcaagagac
     4081 agctggtgga aacccggcag atcacaaagc acgtggcaca gatcctggac tcccggatga
     4141 acactaagta cgacgagaat gacaagctga tccgggaagt gaaagtgatc accctgaagt
     4201 ccaagctggt gtccgatttc cggaaggatt tccagtttta caaagtgcgc gagatcaaca
     4261 actaccacca cgcccacgac gcctacctga acgccgtcgt gggaaccgcc ctgatcaaaa
     4321 agtaccctaa gctggaaagc gagttcgtgt acggcgacta caaggtgtac gacgtgcgga
     4381 agatgatcgc caagagcgag caggaaatcg gcaaggctac cgccaagtac ttcttctaca
     4441 gcaacatcat gaactttttc aagaccgaga ttaccctggc caacggcgag atccggaagc
     4501 ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt gtgggataag ggccgggatt
     4561 ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa tatcgtgaaa aagaccgagg
     4621 tgcagacagg cggcttcagc aaagagtcta tcctgcccaa gaggaacagc gataagctga
     4681 tcgccagaaa gaaggactgg gaccctaaga agtacggcgg cttcgacagc cccaccgtgg
     4741 cctattctgt gctggtggtg gccaaagtgg aaaagggcaa gtccaagaaa ctgaagagtg
     4801 tgaaagagct gctggggatc accatcatgg aaagaagcag cttcgagaag aatcccatcg
     4861 actttctgga agccaagggc tacaaagaag tgaaaaagga cctgatcatc aagctgccta
     4921 agtactccct gttcgagctg gaaaacggcc ggaagagaat gctggcctct gccggcgaac
     4981 tgcagaaggg aaacgaactg gccctgccct ccaaatatgt gaacttcctg tacctggcca
     5041 gccactatga gaagctgaag ggctcccccg aggataatga gcagaaacag ctgtttgtgg
     5101 aacagcacaa gcactacctg gacgagatca tcgagcagat cagcgagttc tccaagagag
     5161 tgatcctggc cgacgctaat ctggacaaag tgctgtccgc ctacaacaag caccgggata
     5221 agcccatcag agagcaggcc gagaatatca tccacctgtt taccctgacc aatctgggag
     5281 cccctgccgc cttcaagtac tttgacacca ccatcgaccg gaagaggtac accagcacca
     5341 aagaggtgct ggacgccacc ctgatccacc agagcatcac cggcctgtac gagacacgga
     5401 tcgacctgtc tcagctggga ggcgacagcc ccaagaagaa gagaaaggtg gaggccagct
     5461 aagaattcct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     5521 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     5581 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     5641 tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc atgctgggga
     5701 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     5761 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     5821 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     5881 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     5941 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     6001 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     6061 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     6121 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     6181 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     6241 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     6301 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     6361 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6421 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     6481 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     6541 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     6601 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     6661 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6721 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     6781 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     6841 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     6901 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     6961 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     7021 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     7081 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     7141 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     7201 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     7261 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     7321 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     7381 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     7441 ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg tatcattgca
     7501 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     7561 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     7621 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     7681 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     7741 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7801 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     7861 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     7921 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     7981 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     8041 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     8101 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     8161 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     8221 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     8281 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     8341 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     8401 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
