LOCUS       sequence-399746-       13590 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector modified to express two sgRNAs that delete the SVA
            within the SVA-lncRNA AK057321 gene (exon 3) with EF-1apha for Cas9 
            and mcherry expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13590)
  AUTHORS   Nadler MJS, Chang W, Ozkaynak E, Huo Y, Nong Y, Boillot M, Johnson 
            M, Moreno A, Matthew P Anderson
  TITLE     Hominoid SVA-lncRNA AK057321 targets human-specific SVA 
            retrotransposons in SCN8A and CDK5RAP2 to initiate neuronal 
            maturation.
  JOURNAL   Commun Biol. 2023 Mar 30;6(1):347. doi: 10.1038/s42003-023-04683-8.
  PUBMED    36997626
REFERENCE   2  (bases 1 to 13590)
  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: "Commun 
            Biol."; date: "2023-03-30"; volume: "6(1):347. doi"; pages: " 
            10.1038/s42003-023-04683-8"
COMMENT     Sequence Label: lentiCRISPRv2-sgRNAs-del-AK057321-SVA-mcherry
FEATURES             Location/Qualifiers
     source          1..13590
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      196..624
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(283..302)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     493..514
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(633..653)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        638..967
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      818..953
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     880..899
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        1015..1062
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1081..1455
                     /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    1585..1706
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1622..1641)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1676..1695
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1755..1771)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1755..1771)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1768..1790)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1779..1795
                     /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(1803..1809,1810..1827,1828..1833))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1848..1869
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1986..2003)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2157..2745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2237..2256)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2916..3707,3708..3776))
                     /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     3539..3558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3777..3881)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3956..3975)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4151..4530
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4532..4730
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4685..4705
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             4748..4928
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4975..5100
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5593..5826
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6011..6055
                     /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    6353..6470
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6521..6769
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        6791..6866
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        6934..7182
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        7204..7279
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7341..7552
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      7571..7580
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7577..11680
                     /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             7577..11680
                     /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             11681..11728
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             11729..11752
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             11762..11818
                     /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"
     regulatory      11855..11864
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             11861..12571
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(12006..12024)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     12274..12293
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    12591..13179
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(12644..12664)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(13062..13073)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             13251..13484
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(13510..13527)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    join(13516..13590,1..150)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
       61 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
      121 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
      181 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
      241 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
      301 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
      361 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
      421 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
      481 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
      541 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
      601 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
      661 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
      721 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
      781 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
      841 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
      901 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
      961 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     1021 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     1081 atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc
     1141 gagttctgga ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt
     1201 gtggtccggg acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac
     1261 aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag
     1321 gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag
     1381 ccgtgggggc gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc
     1441 gaggagcagg actgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     1501 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     1561 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     1621 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     1681 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     1741 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     1801 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     1861 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     1921 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     1981 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     2041 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2101 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     2161 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     2221 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     2281 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     2341 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     2401 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     2461 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     2521 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     2581 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     2641 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     2701 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     2761 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     2821 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     2881 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     2941 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     3001 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     3061 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     3121 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     3181 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     3241 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     3301 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     3361 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     3421 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     3481 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     3541 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     3601 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     3661 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     3721 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     3781 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     3841 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     3901 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     3961 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     4021 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     4081 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
     4141 cgcgcgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     4201 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     4261 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     4321 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     4381 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     4441 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     4501 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     4561 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     4621 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     4681 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc
     4741 ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     4801 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     4861 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     4921 ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct
     4981 cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg
     5041 agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc
     5101 agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga
     5161 aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca
     5221 gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa
     5281 ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc
     5341 tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag
     5401 gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg
     5461 aggaggagat atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat
     5521 tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag
     5581 agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg
     5641 cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca
     5701 gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg
     5761 gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca
     5821 gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa
     5881 tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg
     5941 ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa
     6001 ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa
     6061 ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg
     6121 cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg
     6181 atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga
     6241 aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg
     6301 atcggcactg cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa
     6361 gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag
     6421 acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt
     6481 acagggacag cagagatcca gtttggttaa ttaaggtacc gagggcctat ttcccatgat
     6541 tccttcatat ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac
     6601 tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta
     6661 gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa
     6721 gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg gacagcaatt
     6781 tagcagtatc gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac
     6841 ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc tagctaggtc ttgaaaggag
     6901 tgggaattgg ctccggtgcc cgtcagtctc gaggagggcc tatttcccat gattccttca
     6961 tatttgcata tacgatacaa ggctgttaga gagataatta gaattaattt gactgtaaac
     7021 acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca
     7081 gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc
     7141 gatttcttgg ctttatatat cttgtggaaa ggacgaaaca ccggctttgg atacctcgta
     7201 ggtgttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
     7261 agtggcaccg agtcggtgct tttttgaatt cgctagctag gtcttgaaag gagtgggaat
     7321 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
     7381 gggaggggtc ggcaattgat ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
     7441 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
     7501 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggaccggtt
     7561 ctagagcgct gccaccatgg acaagaagta cagcatcggc ctggacatcg gcaccaactc
     7621 tgtgggctgg gccgtgatca ccgacgagta caaggtgccc agcaagaaat tcaaggtgct
     7681 gggcaacacc gaccggcaca gcatcaagaa gaacctgatc ggagccctgc tgttcgacag
     7741 cggcgaaaca gccgaggcca cccggctgaa gagaaccgcc agaagaagat acaccagacg
     7801 gaagaaccgg atctgctatc tgcaagagat cttcagcaac gagatggcca aggtggacga
     7861 cagcttcttc cacagactgg aagagtcctt cctggtggaa gaggataaga agcacgagcg
     7921 gcaccccatc ttcggcaaca tcgtggacga ggtggcctac cacgagaagt accccaccat
     7981 ctaccacctg agaaagaaac tggtggacag caccgacaag gccgacctgc ggctgatcta
     8041 tctggccctg gcccacatga tcaagttccg gggccacttc ctgatcgagg gcgacctgaa
     8101 ccccgacaac agcgacgtgg acaagctgtt catccagctg gtgcagacct acaaccagct
     8161 gttcgaggaa aaccccatca acgccagcgg cgtggacgcc aaggccatcc tgtctgccag
     8221 actgagcaag agcagacggc tggaaaatct gatcgcccag ctgcccggcg agaagaagaa
     8281 tggcctgttc ggaaacctga ttgccctgag cctgggcctg acccccaact tcaagagcaa
     8341 cttcgacctg gccgaggatg ccaaactgca gctgagcaag gacacctacg acgacgacct
     8401 ggacaacctg ctggcccaga tcggcgacca gtacgccgac ctgtttctgg ccgccaagaa
     8461 cctgtccgac gccatcctgc tgagcgacat cctgagagtg aacaccgaga tcaccaaggc
     8521 ccccctgagc gcctctatga tcaagagata cgacgagcac caccaggacc tgaccctgct
     8581 gaaagctctc gtgcggcagc agctgcctga gaagtacaaa gagattttct tcgaccagag
     8641 caagaacggc tacgccggct acattgacgg cggagccagc caggaagagt tctacaagtt
     8701 catcaagccc atcctggaaa agatggacgg caccgaggaa ctgctcgtga agctgaacag
     8761 agaggacctg ctgcggaagc agcggacctt cgacaacggc agcatccccc accagatcca
     8821 cctgggagag ctgcacgcca ttctgcggcg gcaggaagat ttttacccat tcctgaagga
     8881 caaccgggaa aagatcgaga agatcctgac cttccgcatc ccctactacg tgggccctct
     8941 ggccagggga aacagcagat tcgcctggat gaccagaaag agcgaggaaa ccatcacccc
     9001 ctggaacttc gaggaagtgg tggacaaggg cgcttccgcc cagagcttca tcgagcggat
     9061 gaccaacttc gataagaacc tgcccaacga gaaggtgctg cccaagcaca gcctgctgta
     9121 cgagtacttc accgtgtata acgagctgac caaagtgaaa tacgtgaccg agggaatgag
     9181 aaagcccgcc ttcctgagcg gcgagcagaa aaaggccatc gtggacctgc tgttcaagac
     9241 caaccggaaa gtgaccgtga agcagctgaa agaggactac ttcaagaaaa tcgagtgctt
     9301 cgactccgtg gaaatctccg gcgtggaaga tcggttcaac gcctccctgg gcacatacca
     9361 cgatctgctg aaaattatca aggacaagga cttcctggac aatgaggaaa acgaggacat
     9421 tctggaagat atcgtgctga ccctgacact gtttgaggac agagagatga tcgaggaacg
     9481 gctgaaaacc tatgcccacc tgttcgacga caaagtgatg aagcagctga agcggcggag
     9541 atacaccggc tggggcaggc tgagccggaa gctgatcaac ggcatccggg acaagcagtc
     9601 cggcaagaca atcctggatt tcctgaagtc cgacggcttc gccaacagaa acttcatgca
     9661 gctgatccac gacgacagcc tgacctttaa agaggacatc cagaaagccc aggtgtccgg
     9721 ccagggcgat agcctgcacg agcacattgc caatctggcc ggcagccccg ccattaagaa
     9781 gggcatcctg cagacagtga aggtggtgga cgagctcgtg aaagtgatgg gccggcacaa
     9841 gcccgagaac atcgtgatcg aaatggccag agagaaccag accacccaga agggacagaa
     9901 gaacagccgc gagagaatga agcggatcga agagggcatc aaagagctgg gcagccagat
     9961 cctgaaagaa caccccgtgg aaaacaccca gctgcagaac gagaagctgt acctgtacta
    10021 cctgcagaat gggcgggata tgtacgtgga ccaggaactg gacatcaacc ggctgtccga
    10081 ctacgatgtg gaccatatcg tgcctcagag ctttctgaag gacgactcca tcgacaacaa
    10141 ggtgctgacc agaagcgaca agaaccgggg caagagcgac aacgtgccct ccgaagaggt
    10201 cgtgaagaag atgaagaact actggcggca gctgctgaac gccaagctga ttacccagag
    10261 aaagttcgac aatctgacca aggccgagag aggcggcctg agcgaactgg ataaggccgg
    10321 cttcatcaag agacagctgg tggaaacccg gcagatcaca aagcacgtgg cacagatcct
    10381 ggactcccgg atgaacacta agtacgacga gaatgacaag ctgatccggg aagtgaaagt
    10441 gatcaccctg aagtccaagc tggtgtccga tttccggaag gatttccagt tttacaaagt
    10501 gcgcgagatc aacaactacc accacgccca cgacgcctac ctgaacgccg tcgtgggaac
    10561 cgccctgatc aaaaagtacc ctaagctgga aagcgagttc gtgtacggcg actacaaggt
    10621 gtacgacgtg cggaagatga tcgccaagag cgagcaggaa atcggcaagg ctaccgccaa
    10681 gtacttcttc tacagcaaca tcatgaactt tttcaagacc gagattaccc tggccaacgg
    10741 cgagatccgg aagcggcctc tgatcgagac aaacggcgaa accggggaga tcgtgtggga
    10801 taagggccgg gattttgcca ccgtgcggaa agtgctgagc atgccccaag tgaatatcgt
    10861 gaaaaagacc gaggtgcaga caggcggctt cagcaaagag tctatcctgc ccaagaggaa
    10921 cagcgataag ctgatcgcca gaaagaagga ctgggaccct aagaagtacg gcggcttcga
    10981 cagccccacc gtggcctatt ctgtgctggt ggtggccaaa gtggaaaagg gcaagtccaa
    11041 gaaactgaag agtgtgaaag agctgctggg gatcaccatc atggaaagaa gcagcttcga
    11101 gaagaatccc atcgactttc tggaagccaa gggctacaaa gaagtgaaaa aggacctgat
    11161 catcaagctg cctaagtact ccctgttcga gctggaaaac ggccggaaga gaatgctggc
    11221 ctctgccggc gaactgcaga agggaaacga actggccctg ccctccaaat atgtgaactt
    11281 cctgtacctg gccagccact atgagaagct gaagggctcc cccgaggata atgagcagaa
    11341 acagctgttt gtggaacagc acaagcacta cctggacgag atcatcgagc agatcagcga
    11401 gttctccaag agagtgatcc tggccgacgc taatctggac aaagtgctgt ccgcctacaa
    11461 caagcaccgg gataagccca tcagagagca ggccgagaat atcatccacc tgtttaccct
    11521 gaccaatctg ggagcccctg ccgccttcaa gtactttgac accaccatcg accggaagag
    11581 gtacaccagc accaaagagg tgctggacgc caccctgatc caccagagca tcaccggcct
    11641 gtacgagaca cggatcgacc tgtctcagct gggaggcgac aagcgacctg ccgccacaaa
    11701 gaaggctgga caggctaaga agaagaaaga ttacaaagac gatgacgata agggatccgg
    11761 cgcaacaaac ttctctctgc tgaaacaagc cggagatgtc gaagagaatc ctggaccgaa
    11821 aaagcctgaa ctcaccgcga cgtctgtccc ggtcgccacc atggtgagca agggcgagga
    11881 ggataacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg agggctccgt
    11941 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca
    12001 gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc
    12061 ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca tccccgacta
    12121 cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg
    12181 cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca tctacaaggt
    12241 gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga agaccatggg
    12301 ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa
    12361 gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga ccacctacaa
    12421 ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt tggacatcac
    12481 ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg gccgccactc
    12541 caccggcggc atggacgagc tgtacaagta actagacgcg ttaagtcgac aatcaacctc
    12601 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
    12661 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
    12721 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
    12781 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
    12841 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
    12901 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
    12961 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
    13021 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
    13081 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
    13141 ctcagacgag tcggatctcc ctttgggccg cctccccgcg tcgactttaa gaccaatgac
    13201 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
    13261 aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga
    13321 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
    13381 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
    13441 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc
    13501 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
    13561 tgccttcctt gaccctggaa ggtgccactc
//
