LOCUS       sequence-237636-       15069 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector for the delivery of multiple sgRNAs targeting 
            different genes with constitutive Cas9 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15069)
  AUTHORS   Cao J, Wu L, Zhang SM, Lu M, Cheung WK, Cai W, Gale M, Xu Q, Yan Q
  TITLE     An easy and efficient inducible CRISPR/Cas9 platform with improved 
            specificity for multiple gene targeting.
  JOURNAL   Nucleic Acids Res. 2016 Nov 2;44(19):e149. Epub 2016 Jul 25.
  PUBMED    27458201
REFERENCE   2  (bases 1 to 15069)
  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: "Nucleic 
            Acids Res."; date: "2016-11-2"; volume: "44(19)"; pages: "e149. Epub
            2016 Jul 25"
COMMENT     Sequence Label: Lenti-multi-CRISPR
FEATURES             Location/Qualifiers
     source          1..15069
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     15..34
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        150..197
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             216..590
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    720..841
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(757..776)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     811..830
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(890..906)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(890..906)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(903..925)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    914..930
                     /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(join(938..944,945..962,963..968))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    983..1004
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1121..1138)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1292..1880)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1372..1391)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2051..2842,2843..2911))
                     /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     2674..2693
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2912..3016)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3091..3110)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3282..3661
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3663..3861
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3816..3836
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             3879..4059
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4106..4231
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4724..4957
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5142..5186
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    5484..5601
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5652..5900
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     promoter        join(5924..5929,5930..5947,5948..5954)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             6008..6667
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(6074..6093)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             7009..7314
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     7193..7212
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     misc_RNA        7353..7428
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7721..7969
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        7978..8053
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        8115..8326
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      8345..8354
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8351..12454
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8351..12454
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             12455..12502
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             12503..12526
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             12536..12592
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             12593..13189
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     13086..13106
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    13205..13793
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(13258..13278)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(13676..13687)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             13865..14098
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(14124..14141)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    14130..14354
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      14400..14828
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(14487..14506)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     14697..14718
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(14837..14857)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(14842..15069,1..102)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(15022..15069,1..88)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
       61 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
      121 atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca tagtatatcg
      181 gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc agtgccgttc
      241 cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac cggctcgggt
      301 tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt
      361 tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg gtgtgggtgc
      421 gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac ttccgggacg
      481 cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag ttcgccctgc
      541 gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga cacgtgctac
      601 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
      661 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
      721 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
      781 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
      841 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
      901 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
      961 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     1021 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     1081 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     1141 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     1201 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     1261 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     1321 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     1381 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     1441 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     1501 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     1561 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     1621 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     1681 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     1741 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     1801 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     1861 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     1921 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     1981 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     2041 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     2101 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     2161 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     2221 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     2281 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     2341 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     2401 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     2461 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     2521 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     2581 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     2641 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     2701 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     2761 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     2821 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     2881 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     2941 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     3001 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     3061 atctcccgat cccctatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     3121 ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta
     3181 agctacaaca aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gggttaggcg
     3241 ttttgcgctg cttcgcgatg tacgggccag atatacgcgt tgacattgat tattgactag
     3301 ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt
     3361 tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac
     3421 gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg
     3481 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag
     3541 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat
     3601 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
     3661 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
     3721 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
     3781 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
     3841 gtgggaggtc tatataagca gcgcgttttg cctgtactgg gtctctctgg ttagaccaga
     3901 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
     3961 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
     4021 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt
     4081 gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg
     4141 cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg
     4201 ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg
     4261 atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt
     4321 atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga
     4381 aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat cagaagaact
     4441 tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa
     4501 agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc
     4561 acagcaagcg gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa
     4621 gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg
     4681 caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg
     4741 ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg
     4801 ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg
     4861 cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc
     4921 tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa
     4981 aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac
     5041 agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct
     5101 taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat
     5161 tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt
     5221 atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg
     5281 tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc
     5341 tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag
     5401 acagagacag atccattcga ttagtgaacg gatcggcact gcgtgcgcca attctgcaga
     5461 caaatggcag tattcatcca caattttaaa agaaaagggg ggattggggg gtacagtgca
     5521 ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa
     5581 attacaaaaa ttcaaaattt tcgggtttat tacagggaca gcagagatcc agtttggtta
     5641 attaaggtac cgagggccta tttcccatga ttccttcata tttgcatata cgatacaagg
     5701 ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta gtacaaaata
     5761 cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa
     5821 tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct
     5881 tgtggaaagg acgaaacacc ggagacgatt aggcacccca ggctttacac tttatgcttc
     5941 cggctcgtat aatgtgtgga ttttgagtta ggatccgtcg agattttcag gagctaagga
     6001 agctaaaatg gagaaaaaaa tcactggata taccaccgtt gatatatccc aatggcatcg
     6061 taaagaacat tttgaggcat ttcagtcagt tgctcaatgt acctataacc agaccgttca
     6121 gctggatatt acggcctttt taaagaccgt aaagaaaaat aagcacaagt tttatccggc
     6181 ctttattcac attcttgccc gcctgatgaa tgctcatccg gaattccgta tggcaatgaa
     6241 agacggtgag ctggtgatat gggatagtgt tcacccttgt tacaccgttt tccatgagca
     6301 aactgaaacg ttttcatcgc tctggagtga ataccacgac gatttccggc agtttctaca
     6361 catatattcg caagatgtgg cgtgttacgg tgaaaacctg gcctatttcc ctaaagggtt
     6421 tattgagaat atgtttttcg tctcagccaa tccctgggtg agtttcacca gttttgattt
     6481 aaacgtggcc aatatggaca acttcttcgc ccccgttttc accatgggca aatattatac
     6541 gcaaggcgac aaggtgctga tgccgctggc gattcaggtt catcatgccg tttgtgatgg
     6601 cttccatgtc ggcagaatgc ttaatgaatt acaacagtac tgcgatgagt ggcagggcgg
     6661 ggcgtaaaga tctggatccg gcttactaaa agccagataa cagtatgcgt atttgcgcgc
     6721 tgatttttgc ggtataagaa tatatactga tatgtatacc cgaagtatgt caaaaagagg
     6781 tatgctatga agcagcgtat tacagtgaca gttgacagcg acagctatca gttgctcaag
     6841 gcatatatga tgtcaatatc tccggtctgg taagcacaac catgcagaat gaagcccgtc
     6901 gtctgcgtgc cgaacgctgg aaagcggaaa atcaggaagg gatggctgag gtcgcccggt
     6961 ttattgaaat gaacggctct tttgctgacg agaacagggg ctggtgaaat gcagtttaag
     7021 gtttacacct ataaaagaga gagccgttat cgtctgtttg tggatgtaca gagtgatatt
     7081 attgacacgc ccgggcgacg gatggtgatc cccctggcca gtgcacgtct gctgtcagat
     7141 aaagtctccc gtgaacttta cccggtggtg catatcgggg atgaaagctg gcgcatgatg
     7201 accaccgata tggccagtgt gccggtctcc gttatcgggg aagaagtggc tgatctcagc
     7261 caccgcgaaa atgacatcaa aaacgccatt aacctgatgt tctggggaat ataaatgtca
     7321 ggctccctta tacacagcca gtctgcagct gggttttaga gctagaaata gcaagttaaa
     7381 ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttaagctt
     7441 ggcgtaacta gatcttgaga caaatggcag tattcatcca caattttaaa agaaaagggg
     7501 ggattggggg gtacagtgca ggggaaagag tccggcaagt ttgtggaatt ggtttaacat
     7561 aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt
     7621 aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt
     7681 atcgtttcag acccacctcc caaccccgag gggacccaga gagggcctat ttcccatgat
     7741 tccttcatat ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac
     7801 tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta
     7861 gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa
     7921 gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg cgtctctgtt
     7981 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     8041 accgagtcgg tgcttttttg aattcgctag ctaggtcttg aaaggagtgg gaattggctc
     8101 cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg
     8161 ggtcggcaat tgatccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt
     8221 cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt
     8281 cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggacc ggttctagag
     8341 cgctgccacc atggacaaga agtacagcat cggcctggac atcggcacca actctgtggg
     8401 ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa
     8461 caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga
     8521 aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa
     8581 ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt
     8641 cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc
     8701 catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca
     8761 cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc
     8821 cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga
     8881 caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga
     8941 ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag
     9001 caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct
     9061 gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga
     9121 cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa
     9181 cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc
     9241 cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct
     9301 gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc
     9361 tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa
     9421 cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa
     9481 gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga
     9541 cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg
     9601 agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg
     9661 ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag
     9721 gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa
     9781 cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa
     9841 cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta
     9901 cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc
     9961 cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg
    10021 gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc
    10081 cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct
    10141 gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga
    10201 agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa
    10261 aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac
    10321 cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa
    10381 gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat
    10441 ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg
    10501 cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat
    10561 cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga
    10621 gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag
    10681 ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa
    10741 agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca
    10801 gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga
    10861 tgtggaccat atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct
    10921 gaccagaagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa
    10981 gaagatgaag aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt
    11041 cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat
    11101 caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc
    11161 ccggatgaac actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac
    11221 cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga
    11281 gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct
    11341 gatcaaaaag taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga
    11401 cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt
    11461 cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat
    11521 ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg
    11581 ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa
    11641 gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga
    11701 taagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc
    11761 caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact
    11821 gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa
    11881 tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa
    11941 gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc
    12001 cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta
    12061 cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct
    12121 gtttgtggaa cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc
    12181 caagagagtg atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca
    12241 ccgggataag cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa
    12301 tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac
    12361 cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga
    12421 gacacggatc gacctgtctc agctgggagg cgacaagcga cctgccgcca caaagaaggc
    12481 tggacaggct aagaagaaga aagattacaa agacgatgac gataagggat ccggcgcaac
    12541 aaacttctct ctgctgaaac aagccggaga tgtcgaagag aatcctggac cgaccgagta
    12601 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc
    12661 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga
    12721 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
    12781 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
    12841 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
    12901 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
    12961 cctggccacc gtcggagtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
    13021 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc
    13081 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
    13141 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgaa cgcgttaagt
    13201 cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
    13261 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
    13321 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
    13381 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
    13441 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
    13501 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
    13561 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
    13621 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
    13681 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
    13741 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgtcgact
    13801 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg
    13861 ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc ttgtactggg
    13921 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
    13981 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
    14041 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagg
    14101 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
    14161 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
    14221 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
    14281 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
    14341 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
    14401 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
    14461 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
    14521 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
    14581 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
    14641 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
    14701 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
    14761 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
    14821 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
    14881 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
    14941 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
    15001 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
    15061 ccccatggc
//
