LOCUS       sequence-373294-       15264 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dCas9 effector fusion for manipulating transcription.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15264)
  AUTHORS   Tycko J, Van MV, Aradhana, DelRosso N, Ye H, Yao D, Valbuena R, 
            Vaughan-Jackson A, Xu X, Ludwig C, Spees K, Liu K, Gu M, Khare V, 
            Mukund AX, Suzuki PH, Arana S, Zhang C, Du PP, Ornstein TS, Hess GT,
            Kamber RA, Qi LS, Khalil AS, Bintu L, Bassik MC
  TITLE     Development of compact transcriptional effectors using 
            high-throughput measurements in diverse contexts.
  JOURNAL   Nat Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6.
  PUBMED    39487265
REFERENCE   2  (bases 1 to 15264)
  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: "Nat 
            Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6."
COMMENT     Sequence Label: pJT340 Lenti pEF-dCas9-3XFLAG-KRAB[WSR7EEE] + ...
FEATURES             Location/Qualifiers
     source          1..15264
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(149..253)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(328..347)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        519..898
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        900..1098
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1053..1073
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             1116..1296
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1343..1468
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1961..2194
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2379..2423
                     /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    2721..2838
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2968..4146
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3199..4137
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4094..4114
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4162..4209
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(4210..4233,4234..4236,4237..6792,6793..6795,
                     6796..8310)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             8365..8385
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8401..8466
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8518..8700
                     /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="TLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGYQ
                     LTKPDVILRLEKGEEP"
     CDS             8977..9030
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             9757..9813
                     /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             9814..10212
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    10237..10825
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(10290..10310)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10708..10719)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(10828..10844)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10829..10845)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             11350..11530
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(11556..11573)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    11562..11786
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11832..12260
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11919..11938)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     12129..12150
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(12269..12289)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        12274..12603
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      12454..12589
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     12516..12535
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        12651..12698
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             12717..13091
                     /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    13221..13342
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(13258..13277)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     13312..13331
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(13391..13407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(13391..13407)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(13404..13426)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    13415..13431
                     /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(13439..13445,13446..13463,13464..13469))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13484..13505
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(13622..13639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(13793..14381)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13873..13892)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(14552..15264,1..79,80..148))
                     /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     15175..15194
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
       61 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
      121 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
      181 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
      241 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
      301 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
      361 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
      421 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
      481 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
      541 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
      601 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
      661 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
      721 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
      781 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
      841 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
      901 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
      961 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
     1021 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
     1081 ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct
     1141 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     1201 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     1261 tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa
     1321 agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac
     1381 ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta
     1441 gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg
     1501 ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg
     1561 ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg
     1621 ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag aagaacttag
     1681 atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga
     1741 caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca
     1801 gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg
     1861 aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa
     1921 agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt
     1981 tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca
     2041 gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc
     2101 aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg
     2161 ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac
     2221 tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga
     2281 tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa
     2341 tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg
     2401 aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata
     2461 taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac
     2521 tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc
     2581 caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca
     2641 gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa
     2701 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     2761 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     2821 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt ttggttaatt
     2881 agctagctgc aaagatggat aaagttttaa acagagagga atctttgcag ctaatggacc
     2941 ttctaggtct tgaaaggagt gggaattggc tccggtgccc gtcagtgggc agagcgcaca
     3001 tcgcccacag tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga
     3061 aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag
     3121 ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg
     3181 tttgccgcca gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg cctctttacg
     3241 ggttatggcc cttgcgtgcc ttgaattact tccacctggc tgcagtacgt gattcttgat
     3301 cccgagcttc gggttggaag tgggtgggag agttcgaggc cttgcgctta aggagcccct
     3361 tcgcctcgtg cttgagttga ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg
     3421 tggcaccttc gcgcctgtct cgctgctttc gataagtctc tagccattta aaatttttga
     3481 tgacctgctg cgacgctttt tttctggcaa gatagtcttg taaatgcggg ccaagatctg
     3541 cacactggta tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc
     3601 acatgttcgg cgaggcgggg cctgcgagcg cggccaccga gaatcggacg ggggtagtct
     3661 caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc cgtgtatcgc cccgccctgg
     3721 gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc
     3781 cctgctgcag ggagctcaaa atggaggacg cggcgctcgg gagagcgggc gggtgagtca
     3841 cccacacaaa ggaaaagggc ctttccgtcc tcagccgtcg cttcatgtga ctccacggag
     3901 taccgggcgc cgtccaggca cctcgattag ttctcgagct tttggagtac gtcgtcttta
     3961 ggttgggggg aggggtttta tgcgatggag tttccccaca ctgagtgggt ggagactgaa
     4021 gttaggccag cttggcactt gatgtaattc tccttggaat ttgccctttt tgagtttgga
     4081 tcttggttca ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc atttcaggtg
     4141 tcgtgacgta cggccaccat gaaaaggccg gcggccacga aaaaggccgg ccaggcaaaa
     4201 aagaaaaagg acaagaagta cagcatcggc ctggccatcg gcaccaactc tgtgggctgg
     4261 gccgtgatca ccgacgagta caaggtgccc agcaagaaat tcaaggtgct gggcaacacc
     4321 gaccggcaca gcatcaagaa gaacctgatc ggagccctgc tgttcgacag cggcgaaaca
     4381 gccgaggcca cccggctgaa gagaaccgcc agaagaagat acaccagacg gaagaaccgg
     4441 atctgctatc tgcaagagat cttcagcaac gagatggcca aggtggacga cagcttcttc
     4501 cacagactgg aagagtcctt cctggtggaa gaggataaga agcacgagcg gcaccccatc
     4561 ttcggcaaca tcgtggacga ggtggcctac cacgagaagt accccaccat ctaccacctg
     4621 agaaagaaac tggtggacag caccgacaag gccgacctgc ggctgatcta tctggccctg
     4681 gcccacatga tcaagttccg gggccacttc ctgatcgagg gcgacctgaa ccccgacaac
     4741 agcgacgtgg acaagctgtt catccagctg gtgcagacct acaaccagct gttcgaggaa
     4801 aaccccatca acgccagcgg cgtggacgcc aaggccatcc tgtctgccag actgagcaag
     4861 agcagacggc tggaaaatct gatcgcccag ctgcccggcg agaagaagaa tggcctgttc
     4921 ggcaacctga ttgccctgag cctgggcctg acccccaact tcaagagcaa cttcgacctg
     4981 gccgaggatg ccaaactgca gctgagcaag gacacctacg acgacgacct ggacaacctg
     5041 ctggcccaga tcggcgacca gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac
     5101 gccatcctgc tgagcgacat cctgagagtg aacaccgaga tcaccaaggc ccccctgagc
     5161 gcctctatga tcaagagata cgacgagcac caccaggacc tgaccctgct gaaagctctc
     5221 gtgcggcagc agctgcctga gaagtacaaa gagattttct tcgaccagag caagaacggc
     5281 tacgccggct acattgacgg cggagccagc caggaagagt tctacaagtt catcaagccc
     5341 atcctggaaa agatggacgg caccgaggaa ctgctcgtga agctgaacag agaggacctg
     5401 ctgcggaagc agcggacctt cgacaacggc agcatccccc accagatcca cctgggagag
     5461 ctgcacgcca ttctgcggcg gcaggaagat ttttacccat tcctgaagga caaccgggaa
     5521 aagatcgaga agatcctgac cttccgcatc ccctactacg tgggccctct ggccagggga
     5581 aacagcagat tcgcctggat gaccagaaag agcgaggaaa ccatcacccc ctggaacttc
     5641 gaggaagtgg tggacaaggg cgcttccgcc cagagcttca tcgagcggat gaccaacttc
     5701 gataagaacc tgcccaacga gaaggtgctg cccaagcaca gcctgctgta cgagtacttc
     5761 accgtgtata acgagctgac caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc
     5821 ttcctgagcg gcgagcagaa aaaggccatc gtggacctgc tgttcaagac caaccggaaa
     5881 gtgaccgtga agcagctgaa agaggactac ttcaagaaaa tcgagtgctt cgactccgtg
     5941 gaaatctccg gcgtggaaga tcggttcaac gcctccctgg gcacatacca cgatctgctg
     6001 aaaattatca aggacaagga cttcctggac aatgaggaaa acgaggacat tctggaagat
     6061 atcgtgctga ccctgacact gtttgaggac agagagatga tcgaggaacg gctgaaaacc
     6121 tatgcccacc tgttcgacga caaagtgatg aagcagctga agcggcggag atacaccggc
     6181 tggggcaggc tgagccggaa gctgatcaac ggcatccggg acaagcagtc cggcaagaca
     6241 atcctggatt tcctgaagtc cgacggcttc gccaacagaa acttcatgca gctgatccac
     6301 gacgacagcc tgacctttaa agaggacatc cagaaagccc aggtgtccgg ccagggcgat
     6361 agcctgcacg agcacattgc caatctggcc ggcagccccg ccattaagaa gggcatcctg
     6421 cagacagtga aggtggtgga cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac
     6481 atcgtgatcg aaatggccag agagaaccag accacccaga agggacagaa gaacagccgc
     6541 gagagaatga agcggatcga agagggcatc aaagagctgg gcagccagat cctgaaagaa
     6601 caccccgtgg aaaacaccca gctgcagaac gagaagctgt acctgtacta cctgcagaat
     6661 gggcgggata tgtacgtgga ccaggaactg gacatcaacc ggctgtccga ctacgatgtg
     6721 gaccacatcg tgcctcagag ctttctgaag gacgactcca tcgacaacaa ggtgctgacc
     6781 agaagcgaca aggcccgggg caagagcgac aacgtgccct ccgaagaggt cgtgaagaag
     6841 atgaagaact actggcggca gctgctgaac gccaagctga ttacccagag aaagttcgac
     6901 aatctgacca aggccgagag aggcggcctg agcgaactgg ataaggccgg cttcatcaag
     6961 agacagctgg tggaaacccg gcagatcaca aagcacgtgg cacagatcct ggactcccgg
     7021 atgaacacta agtacgacga gaatgacaag ctgatccggg aagtgaaagt gatcaccctg
     7081 aagtccaagc tggtgtccga tttccggaag gatttccagt tttacaaagt gcgcgagatc
     7141 aacaactacc accacgccca cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc
     7201 aaaaagtacc ctaagctgga aagcgagttc gtgtacggcg actacaaggt gtacgacgtg
     7261 cggaagatga tcgccaagag cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc
     7321 tacagcaaca tcatgaactt tttcaagacc gagattaccc tggccaacgg cgagatccgg
     7381 aagcggcctc tgatcgagac aaacggcgaa accggggaga tcgtgtggga taagggccgg
     7441 gattttgcca ccgtgcggaa agtgctgagc atgccccaag tgaatatcgt gaaaaagacc
     7501 gaggtgcaga caggcggctt cagcaaagag tctatcctgc ccaagaggaa cagcgataag
     7561 ctgatcgcca gaaagaagga ctgggaccct aagaagtacg gcggcttcga cagccccacc
     7621 gtggcctatt ctgtgctggt ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag
     7681 agtgtgaaag agctgctggg gatcaccatc atggaaagaa gcagcttcga gaagaatccc
     7741 atcgactttc tggaagccaa gggctacaaa gaagtgaaaa aggacctgat catcaagctg
     7801 cctaagtact ccctgttcga gctggaaaac ggccggaaga gaatgctggc ctctgccggc
     7861 gaactgcaga agggaaacga actggccctg ccctccaaat atgtgaactt cctgtacctg
     7921 gccagccact atgagaagct gaagggctcc cccgaggata atgagcagaa acagctgttt
     7981 gtggaacagc acaagcacta cctggacgag atcatcgagc agatcagcga gttctccaag
     8041 agagtgatcc tggccgacgc taatctggac aaagtgctgt ccgcctacaa caagcaccgg
     8101 gataagccca tcagagagca ggccgagaat atcatccacc tgtttaccct gaccaatctg
     8161 ggagcccctg ccgccttcaa gtactttgac accaccatcg accggaagag gtacaccagc
     8221 accaaagagg tgctggacgc caccctgatc caccagagca tcaccggcct gtacgagaca
     8281 cggatcgacc tgtctcagct gggaggcgac agcgctggag gaggtggaag cggaggagga
     8341 ggaagcggag gaggaggtag cggacctaag aaaaagagga aggtggcggc cgctggatcc
     8401 gactacaaag accatgacgg tgattataaa gatcatgaca tcgactacaa ggatgacgat
     8461 gacaagggtg gatccggatc aggctccgat gccaagtcct tgaccgctga ggaggagacc
     8521 ctggtcactt tcaaggacgt ttttgtggat ttcacgaggg aagaatggaa actactagac
     8581 acagcacaac aaatagtata cagaaacgta atgctagaaa actacaaaaa cctagtaagc
     8641 ctaggatacc aactaacaaa accagacgta atactaagac tagaaaaagg agaagaacca
     8701 ggaggaggag gtagcggaat caaggaggag cccctggacg actacgacgg cagctcatct
     8761 tcccccgaaa atgatgatct ctttatgatg ccacggattg taaatgttac atccctcgct
     8821 acggaaggcg gattggtcga tatgggtggt tctggaggag gaggtagcgg acctaggttg
     8881 aagaatgttg atagatccac cgctcaacag ttggccgtga ctgtgggaaa tgtcacagtc
     8941 attatcacag attttaagga aaaaaccgga tctggagagg gaaggggaag cctcctaact
     9001 tgcggagatg tcgaggagaa tcctggccca atggttagta agggagagga gttgataaaa
     9061 gagaatatgc acatgaaact gtatatggaa gggacagtgg acaaccatca cttcaagtgc
     9121 acctctgagg gagaaggaaa gccatacgaa ggcacccaga ctatgcgaat taaggtcgtg
     9181 gagggagggc ctctcccctt cgccttcgat attcttgcaa catctttcct ctacgggagc
     9241 aaaaccttca tcaatcacac acaaggcata cctgactttt ttaagcagag ttttccagag
     9301 ggctttacct gggagcgtgt gactacatac gaagatgggg gtgtgctgac ggctacacag
     9361 gatacttcac ttcaggacgg ttgtctgatc tacaacgtca agatcagggg ggtcaatttc
     9421 acctcgaatg gaccagtaat gcaaaaaaag accctgggat gggaagcttt tactgaaact
     9481 ctctatcccg ccgacggcgg cctcgaaggc agaaacgata tggctttgaa gctggttggc
     9541 gggtcccatc taattgccaa tgccaaaacc acatatagga gcaaaaagcc tgcaaagaac
     9601 ctgaagatgc cgggtgtgta ctatgttgac tatagattag agcggatcaa agaggcaaat
     9661 aacgaaacgt acgtggagca gcacgaggta gccgtggcgc gctactgcga tttaccctcc
     9721 aaactgggcc ataaactgaa ctgtacagga agcggagcta ctaacttcag cctgctgaag
     9781 caggctggcg acgtggaaga aaacccagga cctatggcca agcctttgtc tcaagaagaa
     9841 tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc tgaagactac
     9901 agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat
     9961 cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca
    10021 gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc
    10081 tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag
    10141 gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg
    10201 tgggagggct aagaattcga tatcaagctt atcggtaatc aacctctgga ttacaaaatt
    10261 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
    10321 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
    10381 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
    10441 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
    10501 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
    10561 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
    10621 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg
    10681 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
    10741 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
    10801 atctcccttt gggccgcctc cccgcatcga taccgtcgac ctcgagacct agaaaaacat
    10861 ggagcaatca caagtagcaa tacagcagct accaatgctg attgtgcctg gctagaagca
    10921 caagaggagg aggaggtggg ttttccagtc acacctcagg tacctttaag accaatgact
    10981 tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcta
    11041 attcactccc aacgaagaca agatatcctt gatctgtgga tctaccacac acaaggctac
    11101 ttccctgatt ggcagaacta cacaccaggg ccagggatca gatatccact gacctttgga
    11161 tggtgctaca agctagtacc agttgagcaa gagaaggtag aagaagccaa tgaaggagag
    11221 aacacccgct tgttacaccc tgtgagcctg catgggatgg atgacccgga gagagaagta
    11281 ttagagtgga ggtttgacag ccgcctagca tttcatcaca tggcccgaga gctgcatccg
    11341 gactgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
    11401 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
    11461 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
    11521 atctctagca gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
    11581 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
    11641 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
    11701 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
    11761 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
    11821 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
    11881 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
    11941 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
    12001 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
    12061 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
    12121 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
    12181 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
    12241 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
    12301 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
    12361 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
    12421 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
    12481 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
    12541 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
    12601 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt gttgacaatt
    12661 aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaaccatgg
    12721 ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga gcggtcgagt
    12781 tctggaccga ccggctcggg ttctcccggg acttcgtgga ggacgacttc gccggtgtgg
    12841 tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg ccggacaaca
    12901 ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg tcggaggtcg
    12961 tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc gagcagccgt
    13021 gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc gtggccgagg
    13081 agcaggactg acacgtgcta cgagatttcg attccaccgc cgccttctat gaaaggttgg
    13141 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
    13201 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
    13261 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
    13321 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
    13381 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
    13441 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
    13501 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
    13561 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
    13621 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
    13681 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
    13741 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
    13801 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
    13861 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
    13921 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
    13981 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
    14041 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
    14101 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
    14161 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
    14221 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
    14281 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
    14341 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
    14401 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
    14461 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
    14521 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
    14581 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
    14641 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
    14701 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
    14761 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
    14821 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
    14881 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
    14941 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
    15001 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
    15061 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
    15121 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
    15181 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
    15241 aactctcaag gatcttaccg ctgt
//
