LOCUS       sequence-306093-       14128 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of BE3.9max and customizable gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14128)
  AUTHORS   Hanna RE, Hegde M, Fagre CR, DeWeirdt PC, Sangree AK, Szegletes Z, 
            Griffith A, Feeley MN, Sanson KR, Baidi Y, Koblan LW, Liu DR, Neal 
            JT, Doench JG
  TITLE     Massively parallel assessment of human variants with base editor 
            screens.
  JOURNAL   Cell. 2021 Feb 18;184(4):1064-1080.e20. doi: 
            10.1016/j.cell.2021.01.012.
  PUBMED    33606977
REFERENCE   2  (bases 1 to 14128)
  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"; doi: 
            "10.1016/j.cell.2021.01"; journalName: "Cell"; date: "2021-02-18- 
            18"; volume: "184"; issue: "4"; pages: "1064-1080.e20"
COMMENT     Sequence Label: pRDA_256
FEATURES             Location/Qualifiers
     source          1..14128
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..200
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             218..398
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    445..570
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1063..1296
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1481..1525
                     /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    1823..1940
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1991..2239
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1991..2011
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2162..2181
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2261..2336
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2420..2631
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2681..2701
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2702..3385
                     /codon_start=1
                     /product="cytidine deaminase (C to U editing enzyme) from 
                     rat"
                     /label=APOBEC-1
                     /note="can use ssDNA as a substrate (Komor et al., 2016)"
                     /translation="SSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWGG
                     RHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFLS
                     RYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSNE
                     AHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHIL
                     WATGLK"
     CDS             join(3482..3505,3506..3508,3509..7582)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             3482..7582
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7613..7861
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             7904..7924
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7934..7990
                     /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             7991..8587
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     8484..8504
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8603..9191
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8656..8676)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9074..9085)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9263..9496
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9522..9539)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9528..9752
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9798..10226
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9885..9904)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10095..10116
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10235..10255)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10240..10569
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10420..10555
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10482..10501
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10617..10664
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10683..11057
                     /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    11187..11308
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11224..11243)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11278..11297
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11357..11373)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11357..11373)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11370..11392)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11381..11397
                     /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(11405..11411,11412..11429,11430..11435))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11450..11471
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11588..11605)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11759..12347)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11839..11858)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12518..13309,13310..13378))
                     /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     13141..13160
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13379..13483)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(13558..13577)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        13749..14128
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag cgcgttttgc ctgtactggg tctctctggt tagaccagat
      241 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
      301 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
      361 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg
      421 aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc
      481 acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc
      541 tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga
      601 tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta
      661 tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa
      721 ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt
      781 agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa
      841 gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca
      901 cagcaagcgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
      961 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1021 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1081 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1141 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1201 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1261 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1321 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1381 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1441 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1501 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1561 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1621 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1681 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1741 cagagacaga tccattcgat tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac
     1801 aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag
     1861 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
     1921 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggttaa
     1981 ttaaggtacc gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc
     2041 tgttagagag ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac
     2101 gtgacgtaga aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat
     2161 ggactatcat atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt
     2221 gtggaaagga cgaaacaccg gagacgcttt tttcgtctca gtttgagagc tagaaatagc
     2281 aagttcaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgcgctc
     2341 acctattagc ggctaaggtt ttttgaattc gctagctagg tcttgaaagg agtgggaatt
     2401 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     2461 ggaggggtcg gcaattgatc cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     2521 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     2581 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggaccggtgc
     2641 caccatgaaa cggacagccg acggaagcga gttcgagtca ccaaagaaga agcggaaagt
     2701 ctcctcagag actgggcctg tcgccgtcga tccaaccctg cgccgccgga ttgaacctca
     2761 cgagtttgaa gtgttctttg acccccggga gctgagaaag gagacatgcc tgctgtacga
     2821 gatcaactgg ggaggcaggc actccatctg gaggcacacc tctcagaaca caaataagca
     2881 cgtggaggtg aacttcatcg agaagtttac cacagagcgg tacttctgcc ccaataccag
     2941 atgtagcatc acatggtttc tgagctggtc cccttgcgga gagtgtagca gggccatcac
     3001 cgagttcctg tccagatatc cacacgtgac actgtttatc tacatcgcca ggctgtatca
     3061 ccacgcagac ccaaggaata ggcagggcct gcgcgatctg atcagctccg gcgtgaccat
     3121 ccagatcatg acagagcagg agtccggcta ctgctggcgg aacttcgtga attattctcc
     3181 tagcaacgag gcccactggc ctaggtaccc acacctgtgg gtgcgcctgt acgtgctgga
     3241 gctgtattgc atcatcctgg gcctgccccc ttgtctgaat atcctgcgga gaaagcagcc
     3301 ccagctgacc ttctttacaa tcgccctgca gtcttgtcac tatcagaggc tgccacccca
     3361 catcctgtgg gccacaggcc tgaagtctgg aggatctagc ggaggatcct ctggcagcga
     3421 gacaccagga acaagcgagt cagcaacacc agagagcagt ggcggcagca gcggcggcag
     3481 cgacaagaag tacagcatcg gcctggccat cggcaccaac tctgtgggct gggccgtgat
     3541 caccgacgag tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca
     3601 cagcatcaag aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc
     3661 cacccggctg aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta
     3721 tctgcaagag atcttcagca acgagatggc caaggtggac gacagcttct tccacagact
     3781 ggaagagtcc ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa
     3841 catcgtggac gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa
     3901 actggtggac agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat
     3961 gatcaagttc cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt
     4021 ggacaagctg ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat
     4081 caacgccagc ggcgtggacg ccaaggccat cctgtctgcc agactgagca agagcagacg
     4141 gctggaaaat ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggaaacct
     4201 gattgccctg agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga
     4261 tgccaaactg cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca
     4321 gatcggcgac cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct
     4381 gctgagcgac atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat
     4441 gatcaagaga tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca
     4501 gcagctgcct gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg
     4561 ctacattgac ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga
     4621 aaagatggac ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa
     4681 gcagcggacc ttcgacaacg gcagcatccc ccaccagatc cacctgggag agctgcacgc
     4741 cattctgcgg cggcaggaag atttttaccc attcctgaag gacaaccggg aaaagatcga
     4801 gaagatcctg accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag
     4861 attcgcctgg atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt
     4921 ggtggacaag ggcgcttccg cccagagctt catcgagcgg atgaccaact tcgataagaa
     4981 cctgcccaac gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta
     5041 taacgagctg accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag
     5101 cggcgagcag aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt
     5161 gaagcagctg aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc
     5221 cggcgtggaa gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat
     5281 caaggacaag gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct
     5341 gaccctgaca ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca
     5401 cctgttcgac gacaaagtga tgaagcagct gaagcggcgg agatacaccg gctggggcag
     5461 gctgagccgg aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga
     5521 tttcctgaag tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag
     5581 cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca
     5641 cgagcacatt gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt
     5701 gaaggtggtg gacgagctcg tgaaagtgat gggccggcac aagcccgaga acatcgtgat
     5761 cgaaatggcc agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat
     5821 gaagcggatc gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt
     5881 ggaaaacacc cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga
     5941 tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccatat
     6001 cgtgcctcag agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga
     6061 caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa
     6121 ctactggcgg cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac
     6181 caaggccgag agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct
     6241 ggtggaaacc cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac
     6301 taagtacgac gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa
     6361 gctggtgtcc gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta
     6421 ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta
     6481 ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat
     6541 gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa
     6601 catcatgaac tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc
     6661 tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc
     6721 caccgtgcgg aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca
     6781 gacaggcggc ttcagcaaag agtctatcct gcccaagagg aacagcgata agctgatcgc
     6841 cagaaagaag gactgggacc ctaagaagta cggcggcttc gacagcccca ccgtggccta
     6901 ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa
     6961 agagctgctg gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt
     7021 tctggaagcc aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta
     7081 ctccctgttc gagctggaaa acggccggaa gagaatgctg gcctctgccg gcgaactgca
     7141 gaagggaaac gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca
     7201 ctatgagaag ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca
     7261 gcacaagcac tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat
     7321 cctggccgac gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc
     7381 catcagagag caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc
     7441 tgccgccttc aagtactttg acaccaccat cgaccggaag aggtacacca gcaccaaaga
     7501 ggtgctggac gccaccctga tccaccagag catcaccggc ctgtacgaga cacggatcga
     7561 cctgtctcag ctgggaggtg acagcggcgg gagcggcggg agcgggggga gcactaatct
     7621 gagcgacatc attgagaagg agactgggaa acagctggtc attcaggagt ccatcctgat
     7681 gctgcctgag gaggtggagg aagtgatcgg caacaagcca gagtctgaca tcctggtgca
     7741 caccgcctac gacgagtcca cagatgagaa tgtgatgctg ctgacctctg acgcccccga
     7801 gtataagcct tgggccctgg tcatccagga ttctaacggc gagaataaga tcaagatgct
     7861 gagcggaggc tccaaaagaa ccgccgacgg cagcgagttc gagcccaaga agaagaggaa
     7921 agtcagcgct ggcgcaacaa acttctctct gctgaaacaa gccggagatg tcgaagagaa
     7981 tcctggaccg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     8041 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     8101 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     8161 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     8221 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     8281 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     8341 caaggagccc gcgtggttcc tggccaccgt cggagtctcg cccgaccacc agggcaaggg
     8401 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     8461 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     8521 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     8581 tgcctgaacg cgttaagtcg acaatcaacc tctggattac aaaatttgtg aaagattgac
     8641 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     8701 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     8761 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     8821 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     8881 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     8941 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc
     9001 atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     9061 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     9121 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     9181 cgcctccccg cgtcgacttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     9241 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg
     9301 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     9361 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     9421 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     9481 tggaaaatct ctagcagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag
     9541 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     9601 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     9661 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     9721 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     9781 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     9841 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     9901 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     9961 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
    10021 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
    10081 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
    10141 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
    10201 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
    10261 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
    10321 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
    10381 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
    10441 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
    10501 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
    10561 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg
    10621 acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa
    10681 ccatggccaa gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg
    10741 tcgagttctg gaccgaccgg ctcgggttct cccgggactt cgtggaggac gacttcgccg
    10801 gtgtggtccg ggacgacgtg accctgttca tcagcgcggt ccaggaccag gtggtgccgg
    10861 acaacaccct ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg
    10921 aggtcgtgtc cacgaacttc cgggacgcct ccgggccggc catgaccgag atcggcgagc
    10981 agccgtgggg gcgggagttc gccctgcgcg acccggccgg caactgcgtg cacttcgtgg
    11041 ccgaggagca ggactgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa
    11101 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
    11161 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
    11221 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
    11281 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
    11341 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
    11401 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
    11461 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
    11521 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
    11581 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
    11641 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
    11701 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
    11761 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
    11821 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
    11881 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
    11941 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
    12001 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
    12061 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
    12121 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
    12181 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
    12241 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
    12301 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
    12361 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
    12421 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
    12481 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
    12541 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
    12601 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
    12661 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
    12721 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
    12781 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
    12841 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
    12901 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
    12961 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
    13021 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
    13081 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
    13141 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
    13201 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
    13261 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
    13321 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
    13381 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
    13441 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
    13501 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
    13561 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
    13621 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
    13681 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
    13741 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
    13801 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
    13861 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
    13921 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
    13981 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
    14041 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
    14101 ctacgtatta gtcatcgcta ttaccatg
//
