LOCUS       sequence-222675-       13989 bp    DNA     circular SYN 14-APR-2025
DEFINITION  lenti vector encoding dcas9-3xflag with T2A Blastcidin resistance 
            marker (EF1a-NLS-dCas9-3Xflag-T2A-Blast-WPRE).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13989)
  AUTHORS   Liu LD, Huang M, Dai P, Liu T, Fan S, Cheng X, Zhao Y, Yeap LS, Meng
            FL
  TITLE     Intrinsic Nucleotide Preference of Diversifying Base Editors Guides 
            Antibody Ex Vivo Affinity Maturation.
  JOURNAL   Cell Rep. 2018 Oct 23;25(4):884-892.e3. doi: 
            10.1016/j.celrep.2018.09.090.
  PUBMED    30355495
REFERENCE   2  (bases 1 to 13989)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.celrep.2018.09"; journalName: "Cell Rep"; date: 
            "2018-10-23- 23"; volume: "25"; issue: "4"; pages: "884-892.e3"
COMMENT     Sequence Label: pLL7 lenti-dcas9-3xflag-blast
FEATURES             Location/Qualifiers
     source          1..13989
                     /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             8123..8188
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8204..8257
                     /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             8258..8656
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8681..9269
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8734..8754)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9152..9163)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9272..9288)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9273..9289)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9794..9974
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10000..10017)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10006..10230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10276..10704
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10363..10382)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10573..10594
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10713..10733)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10718..11047
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10898..11033
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10960..10979
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11095..11142
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11161..11535
                     /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    11665..11786
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11702..11721)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11756..11775
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11835..11851)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11835..11851)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11848..11870)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11859..11875
                     /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(11883..11889,11890..11907,11908..11913))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11928..11949
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12066..12083)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12237..12825)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12317..12336)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12996..13787,13788..13856))
                     /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     13619..13638
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13857..13961)
                     /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 gctggatcca tggactataa ggaccacgac ggagactaca aggatcatga
     8161 tattgattac aaagacgatg acgataagtg tacaggcagt ggagagggca gaggaagtct
     8221 gctaacatgc ggtgacgtcg aggagaatcc tggcccaatg gccaagcctt tgtctcaaga
     8281 agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga
     8341 ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt
     8401 atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgctgc
     8461 ggcagctggc aacctgactt gtatcgtcgc gatcggaaat gagaacaggg gcatcttgag
     8521 cccctgcgga cggtgccgac aggtgcttct cgatctgcat cctgggatca aagccatagt
     8581 gaaggacagt gatggacagc cgacggcagt tgggattcgt gaattgctgc cctctggtta
     8641 tgtgtgggag ggctaagaat tcgatatcaa gcttatcggt aatcaacctc tggattacaa
     8701 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata
     8761 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc
     8821 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg
     8881 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac
     8941 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat
     9001 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt
     9061 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat
     9121 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc
     9181 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag
     9241 tcggatctcc ctttgggccg cctccccgca tcgataccgt cgacctcgag acctagaaaa
     9301 acatggagca atcacaagta gcaatacagc agctaccaat gctgattgtg cctggctaga
     9361 agcacaagag gaggaggagg tgggttttcc agtcacacct caggtacctt taagaccaat
     9421 gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg
     9481 gctaattcac tcccaacgaa gacaagatat ccttgatctg tggatctacc acacacaagg
     9541 ctacttccct gattggcaga actacacacc agggccaggg atcagatatc cactgacctt
     9601 tggatggtgc tacaagctag taccagttga gcaagagaag gtagaagaag ccaatgaagg
     9661 agagaacacc cgcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga
     9721 agtattagag tggaggtttg acagccgcct agcatttcat cacatggccc gagagctgca
     9781 tccggactgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
     9841 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
     9901 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
     9961 gaaaatctct agcagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
    10021 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
    10081 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
    10141 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
    10201 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
    10261 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
    10321 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
    10381 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
    10441 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
    10501 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
    10561 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
    10621 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
    10681 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
    10741 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
    10801 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
    10861 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
    10921 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
    10981 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
    11041 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
    11101 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
    11161 atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc
    11221 gagttctgga ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt
    11281 gtggtccggg acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac
    11341 aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag
    11401 gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag
    11461 ccgtgggggc gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc
    11521 gaggagcagg actgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
    11581 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
    11641 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
    11701 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
    11761 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
    11821 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
    11881 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
    11941 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
    12001 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
    12061 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
    12121 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
    12181 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
    12241 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
    12301 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
    12361 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
    12421 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
    12481 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
    12541 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
    12601 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
    12661 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
    12721 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
    12781 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
    12841 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
    12901 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
    12961 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
    13021 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
    13081 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
    13141 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
    13201 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
    13261 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
    13321 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
    13381 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
    13441 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
    13501 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
    13561 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
    13621 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
    13681 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
    13741 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
    13801 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
    13861 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
    13921 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
    13981 ccacctgac
//
