pH-ABE8e

LOCUS       pH-ABE8e               16936 bp ds-DNA     circular SYN 05-JUL-2023
DEFINITION  For plant adenine base editing in rice plants or monocotyledons 
            protoplasts.
ACCESSION   .
VERSION     .
KEYWORDS    pH-ABE8e
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 16936)
  AUTHORS   Xue C, Qiu F, Wang Y, Li B, Zhao KT, Chen K, Gao C
  TITLE     Tuning plant phenotypes by precise, graded downregulation of gene 
            expression.
  JOURNAL   Nat Biotechnol. 2023 Mar 9. doi: 10.1038/s41587-023-01707-w.
  PUBMED    36894598
REFERENCE   2  (bases 1 to 16936)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 5, 2023 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..16936
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(578..1207)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     misc_feature    2508..2532
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     primer_bind     2720..2742
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2734..2751
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2735..2751
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2818..3197
                     /label=OsU3 promoter
                     /note="Oryza sativa (rice) snRNA U3 promoter"
     terminator      3305..3324
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     promoter        3337..5329
                     /label=Ubi promoter
                     /note="maize polyubiquitin gene promoter"
     CDS             5940..10040
                     /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"
     RBS             9953..9961
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             10053..10073
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10086..10106
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10119..10139
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(10798..10814)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10798..10814)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10811..10833)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10822..10838
                     /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)."
     promoter        complement(10846..10876)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10891..10912
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        11103..11779
                     /label=CaMV 35S promoter (enhanced)
                     /note="cauliflower mosaic virus 35S promoter with a 
                     duplicated enhancer region"
     primer_bind     11716..11736
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             11846..12871
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRSQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     K"
     polyA_signal    12912..13086
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    13164..13188
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             13613..14407
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      14494..15082
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     14983..15002
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     15236..15253
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    15268..15408
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(15334..15353)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     15494..15516
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     rep_origin      15752..15946
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(16012..149)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
ORIGIN
        1 tcagcggccg cttgccacga ttcaccgggg cgtgcttctg cctcgatgcg ttgccgctgg
       61 gcggcctgcg cggccttcaa cttctccacc aggtcatcac ccagcgccgc gccgatttgt
      121 accgggccgg atggtttgcg accgctcacg ccgattcctc gggcttgggg gttccagtgc
      181 cattgcaggg ccggcaggca acccagccgc ttacgcctgg ccaaccgccc gttcctccac
      241 acatggggca ttccacggcg tcggtgcctg gttgttcttg attttccatg ccgcctcctt
      301 tagccgctaa aattcatcta ctcatttatt catttgctca tttactctgg tagctgcgcg
      361 atgtattcag atagcagctc ggtaatggtc ttgccttggc gtaccgcgta catcttcagc
      421 ttggtgtgat cctccgccgg caactgaaag ttgacccgct tcatggctgg cgtgtctgcc
      481 aggctggcca acgttgcagc cttgctgctg cgtgcgctcg gacggccggc acttagcgtg
      541 tttgtgcttt tgctcatttt ctctttacct cattaactca aatgagtttt gatttaattt
      601 cagcggccag cgcctggacc tcgcgggcag cgtcgccctc gggttctgat tcaagaacgg
      661 ttgtgccggc ggcggcagtg cctgggtagc tcacgcgctg cgtgatacgg gactcaagaa
      721 tgggcagctc gtacccggcc agcgcctcgg caacctcacc gccgatgcgc gtgcctttga
      781 tcgcccgcga cacgacaaag gccgcttgta gccttccatc cgtgacctca atgcgctgct
      841 taaccagctc caccaggtcg gcggtggccc atatgtcgta agggcttggc tgcaccggaa
      901 tcagcacgaa gtcggctgcc ttgatcgcgg acacagccaa gtccgccgcc tggggcgctc
      961 cgtcgatcac tacgaagtcg cgccggccga tggccttcac gtcgcggtca atcgtcgggc
     1021 ggtcgatgcc gacaacggtt agcggttgat cttcccgcac ggccgcccaa tcgcgggcac
     1081 tgccctgggg atcggaatcg actaacagaa catcggcccc ggcgagttgc agggcgcggg
     1141 ctagatgggt tgcgatggtc gtcttgcctg acccgccttt ctggttaagt acagcgataa
     1201 ccttcatgcg ttccccttgc gtatttgttt atttactcat cgcatcatat acgcagcgac
     1261 cgcatgacgc aagctgtttt actcaaatac acatcacctt tttagacggc ggcgctcggt
     1321 ttcttcagcg gccaagctgg ccggccaggc cgccagcttg gcatcagaca aaccggccag
     1381 gatttcatgc agccgcacgg ttgagacgtg cgcgggcggc tcgaacacgt acccggccgc
     1441 gatcatctcc gcctcgatct cttcggtaat gaaaaacggt tcgtcctggc cgtcctggtg
     1501 cggtttcatg cttgttcctc ttggcgttca ttctcggcgg ccgccagggc gtcggcctcg
     1561 gtcaatgcgt cctcacggaa ggcaccgcgc cgcctggcct cggtgggcgt cacttcctcg
     1621 ctgcgctcaa gtgcgcggta cagggtcgag cgatgcacgc caagcagtgc agccgcctct
     1681 ttcacggtgc ggccttcctg gtcgatcagc tcgcgggcgt gcgcgatctg tgccggggtg
     1741 agggtagggc gggggccaaa cttcacgcct cgggccttgg cggcctcgcg cccgctccgg
     1801 gtgcggtcga tgattaggga acgctcgaac tcggcaatgc cggcgaacac ggtcaacacc
     1861 atgcggccgg ccggcgtggt ggtgtcggcc cacggctctg ccaggctacg caggcccgcg
     1921 ccggcctcct ggatgcgctc ggcaatgtcc agtaggtcgc gggtgctgcg ggccaggcgg
     1981 tctagcctgg tcactgtcac aacgtcgcca gggcgtaggt ggtcaagcat cctggccagc
     2041 tccgggcggt cgcgcctggt gccggtgatc ttctcggaaa acagcttggt gcagccggcc
     2101 gcgtgcagtt cggcccgttg gttggtcaag tcctggtcgt cggtgctgac gcgggcatag
     2161 cccagcaggc cagcggcggc gctcttgttc atggcgtaat gtctccggtt ctagtcgcaa
     2221 gtattctact ttatgcgact aaaacacgcg acaagaaaac gccaggaaaa gggcagggcg
     2281 gcagcctgtc gcgtaactta ggacttgtgc gacatgtcgt tttcagaaga cggctgcact
     2341 gaacgtcaga agccgactgc actatagcag cggaggggtt ggatcaaagt actttgatcc
     2401 cgaggggaac cctgtggttg gcatgcacat acaaatggac gaacggataa accttttcac
     2461 gcccttttaa atatccgatt attctaataa acgctctttt ctcttaggtt tacccgccaa
     2521 tatatcctgt caaacactga tagtttaaac tgaaggcggg aaacgacaat ctgatccaag
     2581 ctcaagctgc tctagcattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     2641 cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt
     2701 tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgccaagctt
     2761 agtaattcat ccaggtcacc aagttctagg attttcagaa ctgcaactta ttttatcaag
     2821 gaatctttaa acatacgaac agatcactta aagttcttct gaagcaactt aaagttatca
     2881 ggcatgcatg gatcttggag gaatcagatg tgcagtcagg gaccatagca caagacaggc
     2941 gtcttctact ggtgctacca gcaaatgctg gaagccggga acactgggta cgttggaaac
     3001 cacgtgatgt gaagaagtaa gataaactgt aggagaaaag catttcgtag tgggccatga
     3061 agcctttcag gacatgtatt gcagtatggg ccggcccatt acgcaattgg acgacaacaa
     3121 agactagtat tagtaccacc tcggctatcc acatagatca aagctgattt aaaagagttg
     3181 tgcagatgat ccgtggcgtg agaccctgca gggtctctgt ttaagagcta tgctggaaac
     3241 agcatagcaa gtttaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg
     3301 gtgctttttt tgttttttat gtctaagctt gcatgcctgc agtgcagcgt gacccggtcg
     3361 tgcccctctc tagagataat gagcattgca tgtctaagtt ataaaaaatt accacatatt
     3421 ttttttgtca cacttgtttg aagtgcagtt tatctatctt tatacatata tttaaacttt
     3481 actctacgaa taatataatc tatagtacta caataatatc agtgttttag agaatcatat
     3541 aaatgaacag ttagacatgg tctaaaggac aattgagtat tttgacaaca ggactctaca
     3601 gttttatctt tttagtgtgc atgtgttctc cttttttttt gcaaatagct tcacctatat
     3661 aatacttcat ccattttatt agtacatcca tttagggttt agggttaatg gtttttatag
     3721 actaattttt ttagtacatc tattttattc tattttagcc tctaaattaa gaaaactaaa
     3781 actctatttt agttttttta tttaataatt tagatataaa atagaataaa ataaagtgac
     3841 taaaaattaa acaaataccc tttaagaaat taaaaaaact aaggaaacat ttttcttgtt
     3901 tcgagtagat aatgccagcc tgttaaacgc cgtcgacgag tctaacggac accaaccagc
     3961 gaaccagcag cgtcgcgtcg ggccaagcga agcagacggc acggcatctc tgtcgctgcc
     4021 tctggacccc tctcgagagt tccgctccac cgttggactt gctccgctgt cggcatccag
     4081 aaattgcgtg gcggagcggc agacgtgagc cggcacggca ggcggcctcc tcctcctctc
     4141 acggcaccgg cagctacggg ggattccttt cccaccgctc cttcgctttc ccttcctcgc
     4201 ccgccgtaat aaatagacac cccctccaca ccctctttcc ccaacctcgt gttgttcgga
     4261 gcgcacacac acacaaccag atctccccca aatccacccg tcggcacctc cgcttcaagg
     4321 tacgccgctc gtcctccccc cccccccctc tctaccttct ctagatcggc gttccggtcc
     4381 atggttaggg cccggtagtt ctacttctgt tcatgtttgt gttagatccg tgtttgtgtt
     4441 agatccgtgc tgctagcgtt cgtacacgga tgcgacctgt acgtcagaca cgttctgatt
     4501 gctaacttgc cagtgtttct ctttggggaa tcctgggatg gctctagccg ttccgcagac
     4561 gggatcgatt tcatgatttt ttttgtttcg ttgcataggg tttggtttgc ccttttcctt
     4621 tatttcaata tatgccgtgc acttgtttgt cgggtcatct tttcatgctt ttttttgtct
     4681 tggttgtgat gatgtggtct ggttgggcgg tcgttctaga tcggagtaga attctgtttc
     4741 aaactacctg gtggatttat taattttgga tctgtatgtg tgtgccatac atattcatag
     4801 ttacgaattg aagatgatgg atggaaatat cgatctagga taggtataca tgttgatgcg
     4861 ggttttactg atgcatatac agagatgctt tttgttcgct tggttgtgat gatgtggtgt
     4921 ggttgggcgg tcgttcattc gttctagatc ggagtagaat actgtttcaa actacctggt
     4981 gtatttatta attttggaac tgtatgtgtg tgtcatacat cttcatagtt acgagtttaa
     5041 gatggatgga aatatcgatc taggataggt atacatgttg atgtgggttt tactgatgca
     5101 tatacatgat ggcatatgca gcatctattc atatgctcta accttgagta cctatctatt
     5161 ataataaaca agtatgtttt ataattattt tgatcttgat atacttggat gatggcatat
     5221 gcagcagcta tatgtggatt tttttagccc tgccttcata cgctatttat ttgcttggta
     5281 ctgtttcttt tgtcgatgct caccctgttg tttggtgtta cttctgcagc cctaggatga
     5341 gcgaagttga atttagccat gaatattgga tgcgccatgc acttacactt gcaaagagag
     5401 cccgcgatga acgcgaagtt cctgttggag cagttcttgt tcttaacaac cgcgttatcg
     5461 gagaaggatg gaaccgcgca atcggacttc atgatcctac agcacatgca gaaatcatgg
     5521 cacttcgcca gggaggactt gttatgcaga actatcgcct tatcgatgca acactttatg
     5581 ttacatttga accttgcgtt atgtgcgcag gagcaatgat ccatagccgc atcggacgcg
     5641 ttgtgttcgg agtacgtaac agcaaacgcg gagcagcagg aagccttatg aacgttctta
     5701 actatcctgg aatgaaccat cgcgttgaaa tcacagaagg aatccttgca gatgaatgcg
     5761 cagcacttct ttgcgatttc taccgtatgc ctcgccaggt gttcaatgcg cagaagaagg
     5821 cccaaagcag catcaactca ggtggtagct ccggcggctc gtcaggttcc gaaacacctg
     5881 gaacaagcga aagcgcaaca cctgaatcta gtggaggaag ttctggtggg tcccttaagg
     5941 acaagaagta ctcgatcggc ctcgccattg ggactaactc tgttggctgg gccgtgatca
     6001 ccgacgagta caaggtgccc tcaaagaagt tcaaggtcct gggcaacacc gatcggcatt
     6061 ccatcaagaa gaatctcatt ggcgctctcc tgttcgacag cggcgagacg gctgaggcta
     6121 cgcggctcaa gcgcaccgcc cgcaggcggt acacgcgcag gaagaatcgc atctgctacc
     6181 tgcaggagat tttctccaac gagatggcga aggttgacga ttctttcttc cacaggctgg
     6241 aggagtcatt cctcgtggag gaggataaga agcacgagcg gcatccaatc ttcggcaaca
     6301 ttgtcgacga ggttgcctac cacgagaagt accctacgat ctaccatctg cggaagaagc
     6361 tcgtggactc cacagataag gcggacctcc gcctgatcta cctcgctctg gcccacatga
     6421 ttaagttcag gggccatttc ctgatcgagg gggatctcaa cccggacaat agcgatgttg
     6481 acaagctgtt catccagctc gtgcagacgt acaaccagct cttcgaggag aaccccatta
     6541 atgcgtcagg cgtcgacgcg aaggctatcc tgtccgctag gctctcgaag tctcggcgcc
     6601 tcgagaacct gatcgcccag ctgccgggcg agaagaagaa cggcctgttc gggaatctca
     6661 ttgcgctcag cctggggctc acgcccaact tcaagtcgaa tttcgatctc gctgaggacg
     6721 ccaagctgca gctctccaag gacacatacg acgatgacct ggataacctc ctggcccaga
     6781 tcggcgatca gtacgcggac ctgttcctcg ctgccaagaa tctgtcggac gccatcctcc
     6841 tgtctgatat tctcagggtg aacaccgaga ttacgaaggc tccgctctca gcctccatga
     6901 tcaagcgcta cgacgagcac catcaggatc tgaccctcct gaaggcgctg gtcaggcagc
     6961 agctccccga gaagtacaag gagatcttct tcgatcagtc gaagaacggc tacgctgggt
     7021 acattgacgg cggggcctct caggaggagt tctacaagtt catcaagccg attctggaga
     7081 agatggacgg cacggaggag ctgctggtga agctcaatcg cgaggacctc ctgaggaagc
     7141 agcggacatt cgataacggc agcatcccac accagattca tctcggggag ctgcacgcta
     7201 tcctgaggag gcaggaggac ttctaccctt tcctcaagga taaccgcgag aagatcgaga
     7261 agattctgac tttcaggatc ccgtactacg tcggcccact cgctaggggc aactcccgct
     7321 tcgcttggat gacccgcaag tcagaggaga cgatcacgcc gtggaacttc gaggaggtgg
     7381 tcgacaaggg cgctagcgct cagtcgttca tcgagaggat gacgaatttc gacaagaacc
     7441 tgccaaatga gaaggtgctc cctaagcact cgctcctgta cgagtacttc acagtctaca
     7501 acgagctgac taaggtgaag tatgtgaccg agggcatgag gaagccggct ttcctgtctg
     7561 gggagcagaa gaaggccatc gtggacctcc tgttcaagac caaccggaag gtcacggtta
     7621 agcagctcaa ggaggactac ttcaagaaga ttgagtgctt cgattcggtc gagatctctg
     7681 gcgttgagga ccgcttcaac gcctccctgg ggacctacca cgatctcctg aagatcatta
     7741 aggataagga cttcctggac aacgaggaga atgaggatat cctcgaggac attgtgctga
     7801 cactcactct gttcgaggac cgggagatga tcgaggagcg cctgaagact tacgcccatc
     7861 tcttcgatga caaggtcatg aagcagctca agaggaggag gtacaccggc tgggggaggc
     7921 tgagcaggaa gctcatcaac ggcattcggg acaagcagtc cgggaagacg atcctcgact
     7981 tcctgaagag cgatggcttc gcgaaccgca atttcatgca gctgattcac gatgacagcc
     8041 tcacattcaa ggaggatatc cagaaggctc aggtgagcgg ccagggggac tcgctgcacg
     8101 agcatatcgc gaacctcgct ggctcgccag ctatcaagaa ggggattctg cagaccgtga
     8161 aggttgtgga cgagctggtg aaggtcatgg gcaggcacaa gcctgagaac atcgtcattg
     8221 agatggcccg ggagaatcag accacgcaga agggccagaa gaactcacgc gagaggatga
     8281 agaggatcga ggagggcatt aaggagctgg ggtcccagat cctcaaggag cacccggtgg
     8341 agaacacgca gctgcagaat gagaagctct acctgtacta cctccagaat ggccgcgata
     8401 tgtatgtgga ccaggagctg gatattaaca ggctcagcga ttacgacgtc gatcatatcg
     8461 ttccacagtc attcctgaag gatgactcca ttgacaacaa ggtcctcacc aggtcggaca
     8521 agaaccgggg caagtctgat aatgttcctt cagaggaggt cgttaagaag atgaagaact
     8581 actggcgcca gctcctgaat gccaagctga tcacgcagcg gaagttcgat aacctcacaa
     8641 aggctgagag gggcgggctc tctgagctgg acaaggcggg cttcatcaag aggcagctgg
     8701 tcgagacacg gcagatcact aagcacgttg cgcagattct cgactcacgg atgaacacta
     8761 agtacgatga gaatgacaag ctgatccgcg aggtgaaggt catcaccctg aagtcaaagc
     8821 tcgtctccga cttcaggaag gatttccagt tctacaaggt tcgggagatc aacaattacc
     8881 accatgccca tgacgcgtac ctgaacgcgg tggtcggcac agctctgatc aagaagtacc
     8941 caaagctcga gagcgagttc gtgtacgggg actacaaggt ttacgatgtg aggaagatga
     9001 tcgccaagtc ggagcaggag attggcaagg ctaccgccaa gtacttcttc tactctaaca
     9061 ttatgaattt cttcaagaca gagatcactc tggccaatgg cgagatccgg aagcgccccc
     9121 tcatcgagac gaacggcgag acgggggaga tcgtgtggga caagggcagg gatttcgcga
     9181 ccgtcaggaa ggttctctcc atgccacaag tgaatatcgt caagaagaca gaggtccaga
     9241 ctggcgggtt ctctaaggag tcaattctgc ctaagcggaa cagcgacaag ctcatcgccc
     9301 gcaagaagga ctgggatccg aagaagtacg gcgggttcga cagccccact gtggcctact
     9361 cggtcctggt tgtggcgaag gttgagaagg gcaagtccaa gaagctcaag agcgtgaagg
     9421 agctgctggg gatcacgatt atggagcgct ccagcttcga gaagaacccg atcgatttcc
     9481 tggaggcgaa gggctacaag gaggtgaaga aggacctgat cattaagctc cccaagtact
     9541 cactcttcga gctggagaac ggcaggaagc ggatgctggc ttccgctggc gagctgcaga
     9601 aggggaacga gctggctctg ccgtccaagt atgtgaactt cctctacctg gcctcccact
     9661 acgagaagct caagggcagc cccgaggaca acgagcagaa gcagctgttc gtcgagcagc
     9721 acaagcatta cctcgacgag atcattgagc agatttccga gttctccaag cgcgtgatcc
     9781 tggccgacgc gaatctggat aaggtcctct ccgcgtacaa caagcaccgc gacaagccaa
     9841 tcagggagca ggctgagaat atcattcatc tcttcaccct gacgaacctc ggcgcccctg
     9901 ctgctttcaa gtacttcgac acaactatcg atcgcaagag gtacacaagc actaaggagg
     9961 tcctggacgc gaccctcatc caccagtcga ttaccggcct ctacgagacg cgcatcgacc
    10021 tgtctcagct cgggggcgac tccggcggca gcccaaagaa gaagcggaag gtgtctggag
    10081 gttctcctaa gaaaaagaga aaagtgtccg gcggctcccc gaagaagaag cgcaaggtgt
    10141 aggagctcag agctttcgtt cgtatcatcg gtttcgacaa cgttcgtcaa gttcaatgca
    10201 tcagtttcat tgcgcacaca ccagaatcct actgagtttg agtattatgg cattgggaaa
    10261 actgtttttc ttgtaccatt tgttgtgctt gtaatttact gtgtttttta ttcggttttc
    10321 gctatcgaac tgtgaaatgg aaatggatgg agaagagtta atgaatgata tggtcctttt
    10381 gttcattctc aaattaatat tatttgtttt ttctcttatt tgttgtgtgt tgaatttgaa
    10441 attataagag atatgcaaac attttgtttt gagtaaaaat gtgtcaaatc gtggcctcta
    10501 atgaccgaag ttaatatgag gagtaaaaca cttgtagttg taccattatg cttattcact
    10561 aggcaacaaa tatattttca gacctagaaa agctgcaaat gttactgaat acaagtatgt
    10621 cctcttgtgt tttagacatt tatgaacttt cctttatgta attttccaga atccttgtca
    10681 gattctaatc attgctttat aattatagtt atactcatgg atttgtagtt gagtatgaaa
    10741 atatttttta atgcatttta tgacttgcca attgattgac aacgaattcg taatcatgtc
    10801 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
    10861 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
    10921 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
    10981 ccaacgcgcg gggagaggcg gtttgcgtat tggctagagc agcttgccaa catggtggag
    11041 cacgacactc tcgtctactc caagaatatc aaagatacag tctcagaaga ccaaagggct
    11101 attgagactt ttcaacaaag ggtaatatcg ggaaacctcc tcggattcca ttgcccagct
    11161 atctgtcact tcatcaaaag gacagtagaa aaggaaggtg gcacctacaa atgccatcat
    11221 tgcgataaag gaaaggctat cgttcaagat gcctctgccg acagtggtcc caaagatgga
    11281 cccccaccca cgaggagcat cgtggaaaaa gaagacgttc caaccacgtc ttcaaagcaa
    11341 gtggattgat gtgaacatgg tggagcacga cactctcgtc tactccaaga atatcaaaga
    11401 tacagtctca gaagaccaaa gggctattga gacttttcaa caaagggtaa tatcgggaaa
    11461 cctcctcgga ttccattgcc cagctatctg tcacttcatc aaaaggacag tagaaaagga
    11521 aggtggcacc tacaaatgcc atcattgcga taaaggaaag gctatcgttc aagatgcctc
    11581 tgccgacagt ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga
    11641 cgttccaacc acgtcttcaa agcaagtgga ttgatgtgat atctccactg acgtaaggga
    11701 tgacgcacaa tcccactatc cttcgcaaga cccttcctct atataaggaa gttcatttca
    11761 tttggagagg acacgctgaa atcaccagtc tctctctaca aatctatctc tctcgagctt
    11821 tcgcagatcc ggggggcaat gagatatgaa aaagcctgaa ctcaccgcga cgtctgtcga
    11881 gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga
    11941 agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag
    12001 ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct
    12061 cccgattccg gaagtgcttg acattgggga gtttagcgag agcctgacct attgcatctc
    12121 ccgccgttca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct
    12181 acaaccggtc gcggaggcta tggatgcgat cgctgcggcc gatcttagcc agacgagcgg
    12241 gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg
    12301 cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc
    12361 gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg
    12421 gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac
    12481 agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat
    12541 cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag
    12601 gcatccggag cttgcaggat cgccacgact ccgggcgtat atgctccgca ttggtcttga
    12661 ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg
    12721 atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag
    12781 aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg
    12841 ccccagcact cgtccgaggg caaagaaata gagtagatgc cgaccgggat ctgtcgatcg
    12901 acaagctcga gtttctccat aataatgtgt gagtagttcc cagataaggg aattagggtt
    12961 cctatagggt ttcgctcatg tgttgagcat ataagaaacc cttagtatgt atttgtattt
    13021 gtaaaatact tctatcaata aaatttctaa ttcctaaaac caaaatccag tactaaaatc
    13081 cagatccccc gaattaattc ggcgttaatt cagtacatta aaaacgtccg caatgtgtta
    13141 ttaagttgtc taagcgtcaa tttgtttaca ccacaatata tcctgccacc agccagccaa
    13201 cagctccccg accggcagct cggcacaaaa tcaccactcg atacaggcag cccatcagtc
    13261 cgggacggcg tcagcgggag agccgttgta aggcggcaga ctttgctcat gttaccgatg
    13321 ctattcggaa gaacggcaac taagctgccg ggtttgaaac acggatgatc tcgcggaggg
    13381 tagcatgttg attgtaacga tgacagagcg ttgctgcctg tgatcaccgc ggtttcaaaa
    13441 tcggctccgt cgatactatg ttatacgcca actttgaaaa caactttgaa aaagctgttt
    13501 tctggtattt aaggttttag aatgcaagga acagtgaatt ggagttcgtc ttgttataat
    13561 tagcttcttg gggtatcttt aaatactgta gaaaagagga aggaaataat aaatggctaa
    13621 aatgagaata tcaccggaat tgaaaaaact gatcgaaaaa taccgctgcg taaaagatac
    13681 ggaaggaatg tctcctgcta aggtatataa gctggtggga gaaaatgaaa acctatattt
    13741 aaaaatgacg gacagccggt ataaagggac cacctatgat gtggaacggg aaaaggacat
    13801 gatgctatgg ctggaaggaa agctgcctgt tccaaaggtc ctgcactttg aacggcatga
    13861 tggctggagc aatctgctca tgagtgaggc cgatggcgtc ctttgctcgg aagagtatga
    13921 agatgaacaa agccctgaaa agattatcga gctgtatgcg gagtgcatca ggctctttca
    13981 ctccatcgac atatcggatt gtccctatac gaatagctta gacagccgct tagccgaatt
    14041 ggattactta ctgaataacg atctggccga tgtggattgc gaaaactggg aagaagacac
    14101 tccatttaaa gatccgcgcg agctgtatga ttttttaaag acggaaaagc ccgaagagga
    14161 acttgtcttt tcccacggcg acctgggaga cagcaacatc tttgtgaaag atggcaaagt
    14221 aagtggcttt attgatcttg ggagaagcgg cagggcggac aagtggtatg acattgcctt
    14281 ctgcgtccgg tcgatcaggg aggatatcgg ggaagaacag tatgtcgagc tattttttga
    14341 cttactgggg atcaagcctg attgggagaa aataaaatat tatattttac tggatgaatt
    14401 gttttagtac ctagaatgca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
    14461 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
    14521 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
    14581 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
    14641 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
    14701 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
    14761 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
    14821 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
    14881 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
    14941 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
    15001 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
    15061 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
    15121 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
    15181 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
    15241 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
    15301 cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
    15361 aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg
    15421 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
    15481 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
    15541 gcgaggcagg gtgccttgat gtgggcgccg gcggtcgagt ggcgacggcg cggcttgtcc
    15601 gcgccctggt agattgcctg gccgtaggcc agccattttt gagcggccag cggccgcgat
    15661 aggccgacgc gaagcggcgg ggcgtaggga gcgcagcgac cgaagggtag gcgctttttg
    15721 cagctcttcg gctgtgcgct ggccagacag ttatgcacag gccaggcggg ttttaagagt
    15781 tttaataagt tttaaagagt tttaggcgga aaaatcgcct tttttctctt ttatatcagt
    15841 cacttacatg tgtgaccggt tcccaatgta cggctttggg ttcccaatgt acgggttccg
    15901 gttcccaatg tacggctttg ggttcccaat gtacgtgcta tccacaggaa acagaccttt
    15961 tcgacctttt tcccctgcta gggcaatttg ccctagcatc tgctccgtac attaggaacc
    16021 ggcggatgct tcgccctcga tcaggttgcg gtagcgcatg actaggatcg ggccagcctg
    16081 ccccgcctcc tccttcaaat cgtactccgg caggtcattt gacccgatca gcttgcgcac
    16141 ggtgaaacag aacttcttga actctccggc gctgccactg cgttcgtaga tcgtcttgaa
    16201 caaccatctg gcttctgcct tgcctgcggc gcggcgtgcc aggcggtaga gaaaacggcc
    16261 gatgccggga tcgatcaaaa agtaatcggg gtgaaccgtc agcacgtccg ggttcttgcc
    16321 ttctgtgatc tcgcggtaca tccaatcagc tagctcgatc tcgatgtact ccggccgccc
    16381 ggtttcgctc tttacgatct tgtagcggct aatcaaggct tcaccctcgg ataccgtcac
    16441 caggcggccg ttcttggcct tcttcgtacg ctgcatggca acgtgcgtgg tgtttaaccg
    16501 aatgcaggtt tctaccaggt cgtctttctg ctttccgcca tcggctcgcc ggcagaactt
    16561 gagtacgtcc gcaacgtgtg gacggaacac gcggccgggc ttgtctccct tcccttcccg
    16621 gtatcggttc atggattcgg ttagatggga aaccgccatc agtaccaggt cgtaatccca
    16681 cacactggcc atgccggccg gccctgcgga aacctctacg tgcccgtctg gaagctcgta
    16741 gcggatcacc tcgccagctc gtcggtcacg cttcgacaga cggaaaacgg ccacgtccat
    16801 gatgctgcga ctatcgcggg tgcccacgtc atagagcatc ggaacgaaaa aatctggttg
    16861 ctcgtcgccc ttgggcggct tcctaatcga cggcgcaccg gctgccggcg gttgccggga
    16921 ttctttgcgg attcga
//