LOCUS       sequence-369765-       14934 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 14934)
  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 14934)
  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: pJT259 Lenti
            pEF-dCas9-3XFLAG-VP64-T2A-tagBFP-...
FEATURES             Location/Qualifiers
     source          1..14934
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2687..3865
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2918..3856
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3813..3833
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3881..3928
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(3929..3952,3953..3955,3956..6511,6512..6514,
                     6515..8029)
                     /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             8084..8104
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8120..8185
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8207..8356
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             8366..8419
                     /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             9146..9202
                     /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             9203..9601
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    9626..10214
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9679..9699)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10097..10108)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(10217..10233)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10218..10234)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             10739..10919
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10945..10962)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10951..11175
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11221..11649
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11308..11327)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11518..11539
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11658..11678)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        11663..11992
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11843..11978
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11905..11924
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        12040..12087
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             12106..12480
                     /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    12610..12731
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12647..12666)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12701..12720
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12780..12796)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12780..12796)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12793..12815)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12804..12820
                     /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(12828..12834,12835..12852,12853..12858))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12873..12894
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(13011..13028)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(13182..13770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13262..13281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13941..14732,14733..14801))
                     /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     14564..14583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14802..14906)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta gctagctgca aagatggata aagttttaaa cagagaggaa
     2641 tctttgcagc taatggacct tctaggtctt gaaaggagtg ggaattggct ccggtgcccg
     2701 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     2761 ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     2821 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     2881 cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg
     2941 cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct
     3001 gcagtacgtg attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc
     3061 ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg
     3121 ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct
     3181 agccatttaa aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt
     3241 aaatgcgggc caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg
     3301 ggcccgtgcg tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag
     3361 aatcggacgg gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc
     3421 gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga
     3481 aagatggccg cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg
     3541 agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc
     3601 ttcatgtgac tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt
     3661 ttggagtacg tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac
     3721 tgagtgggtg gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt
     3781 tgcccttttt gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt
     3841 ttttcttcca tttcaggtgt cgtgacgtac ggccaccatg aaaaggccgg cggccacgaa
     3901 aaaggccggc caggcaaaaa agaaaaagga caagaagtac agcatcggcc tggccatcgg
     3961 caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt
     4021 caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg gagccctgct
     4081 gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca gaagaagata
     4141 caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg agatggccaa
     4201 ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag aggataagaa
     4261 gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc acgagaagta
     4321 ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg ccgacctgcg
     4381 gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc tgatcgaggg
     4441 cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg tgcagaccta
     4501 caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca aggccatcct
     4561 gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc tgcccggcga
     4621 gaagaagaat ggcctgttcg gcaacctgat tgccctgagc ctgggcctga cccccaactt
     4681 caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg acacctacga
     4741 cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc tgtttctggc
     4801 cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga acaccgagat
     4861 caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc accaggacct
     4921 gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag agattttctt
     4981 cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc aggaagagtt
     5041 ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac tgctcgtgaa
     5101 gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca gcatccccca
     5161 ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt tttacccatt
     5221 cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc cctactacgt
     5281 gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga gcgaggaaac
     5341 catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc agagcttcat
     5401 cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc ccaagcacag
     5461 cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat acgtgaccga
     5521 gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg tggacctgct
     5581 gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa gaggactact tcaagaaaat
     5641 cgagtgcttc gactccgtgg aaatctccgg cgtggaagat cggttcaacg cctccctggg
     5701 cacataccac gatctgctga aaattatcaa ggacaaggac ttcctggaca atgaggaaaa
     5761 cgaggacatt ctggaagata tcgtgctgac cctgacactg tttgaggaca gagagatgat
     5821 cgaggaacgg ctgaaaacct atgcccacct gttcgacgac aaagtgatga agcagctgaa
     5881 gcggcggaga tacaccggct ggggcaggct gagccggaag ctgatcaacg gcatccggga
     5941 caagcagtcc ggcaagacaa tcctggattt cctgaagtcc gacggcttcg ccaacagaaa
     6001 cttcatgcag ctgatccacg acgacagcct gacctttaaa gaggacatcc agaaagccca
     6061 ggtgtccggc cagggcgata gcctgcacga gcacattgcc aatctggccg gcagccccgc
     6121 cattaagaag ggcatcctgc agacagtgaa ggtggtggac gagctcgtga aagtgatggg
     6181 ccggcacaag cccgagaaca tcgtgatcga aatggccaga gagaaccaga ccacccagaa
     6241 gggacagaag aacagccgcg agagaatgaa gcggatcgaa gagggcatca aagagctggg
     6301 cagccagatc ctgaaagaac accccgtgga aaacacccag ctgcagaacg agaagctgta
     6361 cctgtactac ctgcagaatg ggcgggatat gtacgtggac caggaactgg acatcaaccg
     6421 gctgtccgac tacgatgtgg accacatcgt gcctcagagc tttctgaagg acgactccat
     6481 cgacaacaag gtgctgacca gaagcgacaa ggcccggggc aagagcgaca acgtgccctc
     6541 cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg ccaagctgat
     6601 tacccagaga aagttcgaca atctgaccaa ggccgagaga ggcggcctga gcgaactgga
     6661 taaggccggc ttcatcaaga gacagctggt ggaaacccgg cagatcacaa agcacgtggc
     6721 acagatcctg gactcccgga tgaacactaa gtacgacgag aatgacaagc tgatccggga
     6781 agtgaaagtg atcaccctga agtccaagct ggtgtccgat ttccggaagg atttccagtt
     6841 ttacaaagtg cgcgagatca acaactacca ccacgcccac gacgcctacc tgaacgccgt
     6901 cgtgggaacc gccctgatca aaaagtaccc taagctggaa agcgagttcg tgtacggcga
     6961 ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc
     7021 taccgccaag tacttcttct acagcaacat catgaacttt ttcaagaccg agattaccct
     7081 ggccaacggc gagatccgga agcggcctct gatcgagaca aacggcgaaa ccggggagat
     7141 cgtgtgggat aagggccggg attttgccac cgtgcggaaa gtgctgagca tgccccaagt
     7201 gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc agcaaagagt ctatcctgcc
     7261 caagaggaac agcgataagc tgatcgccag aaagaaggac tgggacccta agaagtacgg
     7321 cggcttcgac agccccaccg tggcctattc tgtgctggtg gtggccaaag tggaaaaggg
     7381 caagtccaag aaactgaaga gtgtgaaaga gctgctgggg atcaccatca tggaaagaag
     7441 cagcttcgag aagaatccca tcgactttct ggaagccaag ggctacaaag aagtgaaaaa
     7501 ggacctgatc atcaagctgc ctaagtactc cctgttcgag ctggaaaacg gccggaagag
     7561 aatgctggcc tctgccggcg aactgcagaa gggaaacgaa ctggccctgc cctccaaata
     7621 tgtgaacttc ctgtacctgg ccagccacta tgagaagctg aagggctccc ccgaggataa
     7681 tgagcagaaa cagctgtttg tggaacagca caagcactac ctggacgaga tcatcgagca
     7741 gatcagcgag ttctccaaga gagtgatcct ggccgacgct aatctggaca aagtgctgtc
     7801 cgcctacaac aagcaccggg ataagcccat cagagagcag gccgagaata tcatccacct
     7861 gtttaccctg accaatctgg gagcccctgc cgccttcaag tactttgaca ccaccatcga
     7921 ccggaagagg tacaccagca ccaaagaggt gctggacgcc accctgatcc accagagcat
     7981 caccggcctg tacgagacac ggatcgacct gtctcagctg ggaggcgaca gcgctggagg
     8041 aggtggaagc ggaggaggag gaagcggagg aggaggtagc ggacctaaga aaaagaggaa
     8101 ggtggcggcc gctggatccg actacaaaga ccatgacggt gattataaag atcatgacat
     8161 cgactacaag gatgacgatg acaagggtgg atccggatca ggctccgacg cattggacga
     8221 ttttgatctg gatatgctgg gaagtgacgc cctcgatgat tttgaccttg acatgcttgg
     8281 ttcggatgcc cttgatgact ttgacctcga catgctcggc agtgacgccc ttgatgattt
     8341 cgacctggac atgctgggat ctggagaggg aaggggaagc ctcctaactt gcggagatgt
     8401 cgaggagaat cctggcccaa tggttagtaa gggagaggag ttgataaaag agaatatgca
     8461 catgaaactg tatatggaag ggacagtgga caaccatcac ttcaagtgca cctctgaggg
     8521 agaaggaaag ccatacgaag gcacccagac tatgcgaatt aaggtcgtgg agggagggcc
     8581 tctccccttc gccttcgata ttcttgcaac atctttcctc tacgggagca aaaccttcat
     8641 caatcacaca caaggcatac ctgacttttt taagcagagt tttccagagg gctttacctg
     8701 ggagcgtgtg actacatacg aagatggggg tgtgctgacg gctacacagg atacttcact
     8761 tcaggacggt tgtctgatct acaacgtcaa gatcaggggg gtcaatttca cctcgaatgg
     8821 accagtaatg caaaaaaaga ccctgggatg ggaagctttt actgaaactc tctatcccgc
     8881 cgacggcggc ctcgaaggca gaaacgatat ggctttgaag ctggttggcg ggtcccatct
     8941 aattgccaat gccaaaacca catataggag caaaaagcct gcaaagaacc tgaagatgcc
     9001 gggtgtgtac tatgttgact atagattaga gcggatcaaa gaggcaaata acgaaacgta
     9061 cgtggagcag cacgaggtag ccgtggcgcg ctactgcgat ttaccctcca aactgggcca
     9121 taaactgaac tgtacaggaa gcggagctac taacttcagc ctgctgaagc aggctggcga
     9181 cgtggaagaa aacccaggac ctatggccaa gcctttgtct caagaagaat ccaccctcat
     9241 tgaaagagca acggctacaa tcaacagcat ccccatctct gaagactaca gcgtcgccag
     9301 cgcagctctc tctagcgacg gccgcatctt cactggtgtc aatgtatatc attttactgg
     9361 gggaccttgt gcagaactcg tggtgctggg cactgctgct gctgcggcag ctggcaacct
     9421 gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc ttgagcccct gcggacggtg
     9481 ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc atagtgaagg acagtgatgg
     9541 acagccgacg gcagttggga ttcgtgaatt gctgccctct ggttatgtgt gggagggcta
     9601 agaattcgat atcaagctta tcggtaatca acctctggat tacaaaattt gtgaaagatt
     9661 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     9721 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     9781 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     9841 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     9901 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     9961 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
    10021 atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc
    10081 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
    10141 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
    10201 ggccgcctcc ccgcatcgat accgtcgacc tcgagaccta gaaaaacatg gagcaatcac
    10261 aagtagcaat acagcagcta ccaatgctga ttgtgcctgg ctagaagcac aagaggagga
    10321 ggaggtgggt tttccagtca cacctcaggt acctttaaga ccaatgactt acaaggcagc
    10381 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
    10441 acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact tccctgattg
    10501 gcagaactac acaccagggc cagggatcag atatccactg acctttggat ggtgctacaa
    10561 gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga acacccgctt
    10621 gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat tagagtggag
    10681 gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg actgtactgg
    10741 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
    10801 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
    10861 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
    10921 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
    10981 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
    11041 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
    11101 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
    11161 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
    11221 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
    11281 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
    11341 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
    11401 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
    11461 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
    11521 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
    11581 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
    11641 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
    11701 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
    11761 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
    11821 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
    11881 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
    11941 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
    12001 gggagcttgt atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca
    12061 tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc
    12121 agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac
    12181 cggctcgggt tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac
    12241 gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg
    12301 gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac
    12361 ttccgggacg cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag
    12421 ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga
    12481 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
    12541 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
    12601 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
    12661 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
    12721 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
    12781 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
    12841 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
    12901 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
    12961 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
    13021 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
    13081 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
    13141 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
    13201 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
    13261 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
    13321 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
    13381 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
    13441 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
    13501 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
    13561 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
    13621 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
    13681 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
    13741 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
    13801 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
    13861 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
    13921 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
    13981 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
    14041 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
    14101 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
    14161 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
    14221 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
    14281 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
    14341 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
    14401 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
    14461 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
    14521 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
    14581 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
    14641 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
    14701 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
    14761 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
    14821 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
    14881 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgac
//
