LOCUS       sequence-437839-       14394 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral construct for expression of the dCas9-KRAB-MeCP2 CRISPRi 
            vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14394)
  AUTHORS   Pulice JL, Meyerson M
  TITLE     Dosage amplification dictates oncogenic regulation by the NKX2-1 
            lineage factor in lung adenocarcinoma.
  JOURNAL   bioRxiv [Preprint]. 2023 Oct 27:2023.10.26.563996. doi: 
            10.1101/2023.10.26.563996.
  PUBMED    37994369
REFERENCE   2  (bases 1 to 14394)
  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: "bioRxiv 
            [Preprint]. 2023 Oct 27:2023.10.26.563996. doi: 
            10.1101/2023.10.26.563996."
COMMENT     Sequence Label: pLenti_dCas9-KRAB-MeCP2_Blast
FEATURES             Location/Qualifiers
     source          1..14394
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        124..503
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        505..703
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             721..901
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    948..1073
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1566..1799
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1984..2028
                     /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    2326..2443
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2493..2872
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2873..3076
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3073..3097
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      3185..3194
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3191..3217,3218..3220,3221..5704,5705..5710,5711..5776,
                     5777..5779,5780..7294)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=Cas9m4
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D10A, D839A, 
                     H840A, and N863A (Mali et al., 2013)"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVAAIVPQSFLKDDSIDNKVLTRSDKARGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7307..7327
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7388..7582
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             7616..7636
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7646..8503
                     /codon_start=1
                     /product="transcriptional repression domain from rat 
                     methyl-CpG-binding protein 2 (Nan et al., 1997)"
                     /label=MeCP2
                     /translation="VQVKRVLEKSPGKLLVKMPFQASPGGKGEGGGATTSAQVMVIKRP
                     GRKRKAEADPQAIPKKRGRKPGSVVAAAAAEAKKKAVKESSIRSVQETVLPIKKRKTRE
                     TVSIEVKEVVKPLLVSTLGEKSGKGLKTCKSPGRKSKESSPKGRSSSASSPPKKEHHHH
                     HHHAESPKAPMPLLPPPPPPEPQSSEDPISPPEPQDLSSSICKEEKMPRAGSLESDGCP
                     KEPAKTQPMVAAAATTTTTTTTTVAEKYKHRGEGERKDIVSSSMPRPNREEPVDSRTPV
                     TERVS"
     CDS             8513..8569
                     /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             8570..8968
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8993..9581
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9046..9066)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9464..9475)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9584..9600)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9585..9601)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             10106..10286
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10312..10329)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10318..10542
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10588..11016
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10675..10694)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10885..10906
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11025..11045)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      11190..11324
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11251..11270
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11386..11433
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11452..11826
                     /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    11956..12077
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11993..12012)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12047..12066
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12126..12142)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12126..12142)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12139..12161)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12150..12166
                     /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(12174..12180,12181..12198,12199..12204))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12219..12240
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12357..12374)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12528..13116)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12608..12627)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13287..14078,14079..14147))
                     /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     13910..13929
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14148..14252)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(14327..14346)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
       61 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
      121 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
      181 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      241 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      301 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      361 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      421 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      481 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
      541 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
      601 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
      661 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagcgcgttt tgcctgtact
      721 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
      781 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
      841 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
      901 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
      961 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
     1021 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
     1081 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
     1141 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
     1201 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
     1261 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
     1321 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
     1381 aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga
     1441 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
     1501 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
     1561 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
     1621 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
     1681 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1741 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1801 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     1861 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     1921 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     1981 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     2041 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     2101 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     2161 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
     2221 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatcggca
     2281 ctgcgtgcgc caattctgca gacaaatggc agtattcatc cacaatttta aaagaaaagg
     2341 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     2401 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     2461 cagcagagat ccagtttggt taattagcta gcgacattga ttattgacta gttattaata
     2521 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     2581 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     2641 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     2701 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
     2761 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
     2821 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
     2881 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     2941 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     3001 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
     3061 ctatataagc agagctcgtt tagtgaaccg tcagatcgcc tggagacgcc atccacgctg
     3121 ttttgacctc catagaagac accgggaccg atccagcctc cggactctag aggatcgaac
     3181 ccttgccacc atggacaaga agtactccat tgggctcgct atcggcacaa acagcgtcgg
     3241 ctgggccgtc attacggacg agtacaaggt gccgagcaaa aaattcaaag ttctgggcaa
     3301 taccgatcgc cacagcataa agaagaacct cattggcgcc ctcctgttcg actccgggga
     3361 gacggccgaa gccacgcggc tcaaaagaac agcacggcgc agatataccc gcagaaagaa
     3421 tcggatctgc tacctgcagg agatctttag taatgagatg gctaaggtgg atgactcttt
     3481 cttccatagg ctggaggagt cctttttggt ggaggaggat aaaaagcacg agcgccaccc
     3541 aatctttggc aatatcgtgg acgaggtggc gtaccatgaa aagtacccaa ccatatatca
     3601 tctgaggaag aagcttgtag acagtactga taaggctgac ttgcggttga tctatctcgc
     3661 gctggcgcat atgatcaaat ttcggggaca cttcctcatc gagggggacc tgaacccaga
     3721 caacagcgat gtcgacaaac tctttatcca actggttcag acttacaatc agcttttcga
     3781 agagaacccg atcaacgcat ccggagttga cgccaaagca atcctgagcg ctaggctgtc
     3841 caaatcccgg cggctcgaaa acctcatcgc acagctccct ggggagaaga agaacggcct
     3901 gtttggtaat cttatcgccc tgtcactcgg gctgaccccc aactttaaat ctaacttcga
     3961 cctggccgaa gatgccaagc ttcaactgag caaagacacc tacgatgatg atctcgacaa
     4021 tctgctggcc cagatcggcg accagtacgc agaccttttt ttggcggcaa agaacctgtc
     4081 agacgccatt ctgctgagtg atattctgcg agtgaacacg gagatcacca aagctccgct
     4141 gagcgctagt atgatcaagc gctatgatga gcaccaccaa gacttgactt tgctgaaggc
     4201 ccttgtcaga cagcaactgc ctgagaagta caaggaaatt ttcttcgatc agtctaaaaa
     4261 tggctacgcc ggatacattg acggcggagc aagccaggag gaattttaca aatttattaa
     4321 gcccatcttg gaaaaaatgg acggcaccga ggagctgctg gtaaagctta acagagaaga
     4381 tctgttgcgc aaacagcgca ctttcgacaa tggaagcatc ccccaccaga ttcacctggg
     4441 cgaactgcac gctatcctca ggcggcaaga ggatttctac ccctttttga aagataacag
     4501 ggaaaagatt gagaaaatcc tcacatttcg gataccctac tatgtaggcc ccctcgcccg
     4561 gggaaattcc agattcgcgt ggatgactcg caaatcagaa gagaccatca ctccctggaa
     4621 cttcgaggaa gtcgtggata agggggcctc tgcccagtcc ttcatcgaaa ggatgactaa
     4681 ctttgataaa aatctgccta acgaaaaggt gcttcctaaa cactctctgc tgtacgagta
     4741 cttcacagtt tataacgagc tcaccaaggt caaatacgtc acagaaggga tgagaaagcc
     4801 agcattcctg tctggagagc agaagaaagc tatcgtggac ctcctcttca agacgaaccg
     4861 gaaagttacc gtgaaacagc tcaaagaaga ctatttcaaa aagattgaat gtttcgactc
     4921 tgttgaaatc agcggagtgg aggatcgctt caacgcatcc ctgggaacgt atcacgatct
     4981 cctgaaaatc attaaagaca aggacttcct ggacaatgag gagaacgagg acattcttga
     5041 ggacattgtc ctcaccctta cgttgtttga agatagggag atgattgaag aacgcttgaa
     5101 aacttacgct catctcttcg acgacaaagt catgaaacag ctcaagaggc gccgatatac
     5161 aggatggggg cggctgtcaa gaaaactgat caatgggatc cgagacaagc agagtggaaa
     5221 gacaatcctg gattttctta agtccgatgg atttgccaac cggaacttca tgcagttgat
     5281 ccatgatgac tctctcacct ttaaggagga catccagaaa gcacaagttt ctggccaggg
     5341 ggacagtctt cacgagcaca tcgctaatct tgcaggtagc ccagctatca aaaagggaat
     5401 actgcagacc gttaaggtcg tggatgaact cgtcaaagta atgggaaggc ataagcccga
     5461 gaatatcgtt atcgagatgg cccgagagaa ccaaactacc cagaagggac agaagaacag
     5521 tagggaaagg atgaagagga ttgaagaggg tataaaagaa ctggggtccc aaatccttaa
     5581 ggaacaccca gttgaaaaca cccagcttca gaatgagaag ctctacctgt actacctgca
     5641 gaacggcagg gacatgtacg tggatcagga actggacatc aatcggctct ccgactacga
     5701 cgtggctgct atcgtgcccc agtcttttct caaagatgat tctattgata ataaagtgtt
     5761 gacaagatcc gataaagcta gagggaagag tgataacgtc ccctcagaag aagttgtcaa
     5821 gaaaatgaaa aattattggc ggcagctgct gaacgccaaa ctgatcacac aacggaagtt
     5881 cgataatctg actaaggctg aacgaggtgg cctgtctgag ttggataaag ccggcttcat
     5941 caaaaggcag cttgttgaga cacgccagat caccaagcac gtggcccaaa ttctcgattc
     6001 acgcatgaac accaagtacg atgaaaatga caaactgatt cgagaggtga aagttattac
     6061 tctgaagtct aagctggtct cagatttcag aaaggacttt cagttttata aggtgagaga
     6121 gatcaacaat taccaccatg cgcatgatgc ctacctgaat gcagtggtag gcactgcact
     6181 tatcaaaaaa tatcccaagc ttgaatctga atttgtttac ggagactata aagtgtacga
     6241 tgttaggaaa atgatcgcaa agtctgagca ggaaataggc aaggccaccg ctaagtactt
     6301 cttttacagc aatattatga attttttcaa gaccgagatt acactggcca atggagagat
     6361 tcggaagcga ccacttatcg aaacaaacgg agaaacagga gaaatcgtgt gggacaaggg
     6421 tagggatttc gcgacagtcc ggaaggtcct gtccatgccg caggtgaaca tcgttaaaaa
     6481 gaccgaagta cagaccggag gcttctccaa ggaaagtatc ctcccgaaaa ggaacagcga
     6541 caagctgatc gcacgcaaaa aagattggga ccccaagaaa tacggcggat tcgattctcc
     6601 tacagtcgct tacagtgtac tggttgtggc caaagtggag aaagggaagt ctaaaaaact
     6661 caaaagcgtc aaggaactgc tgggcatcac aatcatggag cgatcaagct tcgaaaaaaa
     6721 ccccatcgac tttctcgagg cgaaaggata taaagaggtc aaaaaagacc tcatcattaa
     6781 gcttcccaag tactctctct ttgagcttga aaacggccgg aaacgaatgc tcgctagtgc
     6841 gggcgagctg cagaaaggta acgagctggc actgccctct aaatacgtta atttcttgta
     6901 tctggccagc cactatgaaa agctcaaagg gtctcccgaa gataatgagc agaagcagct
     6961 gttcgtggaa caacacaaac actaccttga tgagatcatc gagcaaataa gcgaattctc
     7021 caaaagagtg atcctcgccg acgctaacct cgataaggtg ctttctgctt acaataagca
     7081 cagggataag cccatcaggg agcaggcaga aaacattatc cacttgttta ctctgaccaa
     7141 cttgggcgcg cctgcagcct tcaagtactt cgacaccacc atagacagaa agcggtacac
     7201 ctctacaaag gaggtcctgg acgccacact gattcatcag tcaattacgg ggctctatga
     7261 aacaagaatc gacctctctc agctcggtgg agacagcagg gctgacccca agaagaagag
     7321 gaaggtgagt ggtggaggaa gtggcgggtc agggtcgatg gacgcgaaat cacttacggc
     7381 atggtcgaga acactggtta cgttcaagga cgtgtttgtg gactttacac gtgaggagtg
     7441 gaaattgctg gatactgcgc aacaaattgt gtatcgaaat gtcatgcttg agaattacaa
     7501 gaacctcgtc agtctcggat accagttgac gaaaccggat gtgatcctta ggctcgaaaa
     7561 gggggaagaa ccttggctgg tatcgggagg tggttcgggt ggctctggat caagcccaaa
     7621 gaagaaacgg aaggtggaag cctcagtgca ggtgaaaagg gtgctggaaa aatcccccgg
     7681 caaactcctc gtgaagatgc ccttccaggc ttcccctggc ggaaaaggtg aagggggtgg
     7741 cgcaaccaca tctgcccagg tcatggtcat caagcgacct ggaaggaaaa gaaaggccga
     7801 ggctgaccct caggccattc caaagaaacg gggacgcaag ccagggtccg tggtcgcagc
     7861 tgcagcagct gaggctaaga aaaaggcagt gaaggaaagc tccatccgca gtgtgcagga
     7921 gactgtcctg cccatcaaga agaggaagac tagggagacc gtgtccatcg aggtcaaaga
     7981 agtggtcaag cccctgctcg tgtccaccct gggcgaaaaa tctggaaagg ggctcaaaac
     8041 atgcaagtca cctggacgga aaagcaagga gtctagtcca aaggggcgct caagctccgc
     8101 ttctagtccc cctaaaaagg aacaccatca ccatcaccat cacgccgagt ctcctaaggc
     8161 tcctatgcca ctgctcccac cacctccacc acctgagcca cagtcaagcg aagaccccat
     8221 cagcccaccc gagcctcagg atctgtcctc tagtatttgc aaagaggaaa agatgcccag
     8281 agcaggcagc ctggagagtg atggctgtcc aaaagaaccc gccaagaccc agcctatggt
     8341 ggcagccgct gcaactacca ccacaaccac aactaccaca gtggccgaaa aatacaagca
     8401 tcgcggcgag ggcgaacgaa aggacattgt gtcaagctcc atgcccagac ctaaccggga
     8461 ggaaccagtc gatagtagga cacccgtgac tgagagagtc tcaggatccg gcgcaacaaa
     8521 cttctctctg ctgaaacaag ccggagatgt cgaagagaat cctggaccga tggccaagcc
     8581 tttgtctcaa gaagaatcca ccctcattga aagagcaacg gctacaatca acagcatccc
     8641 catctctgaa gactacagcg tcgccagcgc agctctctct agcgacggcc gcatcttcac
     8701 tggtgtcaat gtatatcatt ttactggggg accttgtgca gaactcgtgg tgctgggcac
     8761 tgctgctgct gcggcagctg gcaacctgac ttgtatcgtc gcgatcggaa atgagaacag
     8821 gggcatcttg agcccctgcg gacggtgccg acaggtgctt ctcgatctgc atcctgggat
     8881 caaagccata gtgaaggaca gtgatggaca gccgacggca gttgggattc gtgaattgct
     8941 gccctctggt tatgtgtggg agggctaaga attcgatatc aagcttatcg gtaatcaacc
     9001 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     9061 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     9121 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     9181 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     9241 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     9301 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     9361 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt
     9421 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     9481 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     9541 ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc gtcgacctcg
     9601 agacctagaa aaacatggag caatcacaag tagcaataca gcagctacca atgctgattg
     9661 tgcctggcta gaagcacaag aggaggagga ggtgggtttt ccagtcacac ctcaggtacc
     9721 tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg
     9781 gggactggaa gggctaattc actcccaacg aagacaagat atccttgatc tgtggatcta
     9841 ccacacacaa ggctacttcc ctgattggca gaactacaca ccagggccag ggatcagata
     9901 tccactgacc tttggatggt gctacaagct agtaccagtt gagcaagaga aggtagaaga
     9961 agccaatgaa ggagagaaca cccgcttgtt acaccctgtg agcctgcatg ggatggatga
    10021 cccggagaga gaagtattag agtggaggtt tgacagccgc ctagcatttc atcacatggc
    10081 ccgagagctg catccggact gtactgggtc tctctggtta gaccagatct gagcctggga
    10141 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
    10201 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
    10261 ttagtcagtg tggaaaatct ctagcagggc ccgtttaaac ccgctgatca gcctcgactg
    10321 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
    10381 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
    10441 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
    10501 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
    10561 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
    10621 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
    10681 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
    10741 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
    10801 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
    10861 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
    10921 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
    10981 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
    11041 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
    11101 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
    11161 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca gttccgccca
    11221 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
    11281 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
    11341 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
    11401 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag
    11461 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
    11521 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
    11581 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
    11641 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
    11701 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
    11761 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
    11821 gactgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
    11881 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
    11941 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
    12001 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
    12061 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
    12121 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
    12181 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
    12241 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
    12301 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
    12361 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
    12421 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
    12481 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
    12541 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
    12601 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
    12661 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
    12721 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
    12781 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
    12841 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
    12901 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
    12961 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
    13021 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
    13081 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
    13141 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
    13201 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
    13261 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
    13321 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
    13381 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
    13441 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
    13501 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
    13561 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
    13621 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
    13681 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
    13741 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
    13801 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
    13861 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
    13921 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
    13981 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
    14041 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
    14101 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
    14161 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
    14221 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
    14281 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
    14341 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctg
//
