LOCUS       sequence-352550-       13663 bp    DNA     circular SYN 14-APR-2025
DEFINITION  BE Cas expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13663)
  AUTHORS   Sangree AK, Griffith AL, Szegletes ZM, Roy P, DeWeirdt PC, Hegde M, 
            McGee AV, Hanna RE, Doench JG
  TITLE     Benchmarking of SpCas9 variants enables deeper base editor screens 
            of BRCA1 and BCL2.
  JOURNAL   Nat Commun. 2022 Mar 14;13(1):1318. doi: 10.1038/s41467-022-28884-7.
  PUBMED    35288574
REFERENCE   2  (bases 1 to 13663)
  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"; journalName: "Nat 
            Commun."; date: "2022-03-14"; volume: "13(1):1318. doi"; pages: " 
            10.1038/s41467-022-28884-7"
COMMENT     Sequence Label: pRDA_479
FEATURES             Location/Qualifiers
     source          1..13663
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        42..240
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     195..215
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             258..438
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    485..610
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1103..1336
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1521..1565
                     /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    1863..1980
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2031..2279
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2031..2051
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2202..2221
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2301..2376
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2460..2671
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2721..2741
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3336..7436
                     /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
                     SILPKRNSDKLIARKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAKQLQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKQYRSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7479..7499
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7509..7565
                     /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             7566..8162
                     /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     8059..8079
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8178..8766
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8231..8251)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8649..8660)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8838..9071
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9097..9114)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9103..9327
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9373..9801
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9460..9479)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9670..9691
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9810..9830)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9815..10144
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9995..10130
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10057..10076
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10192..10239
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10258..10632
                     /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    10762..10883
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10799..10818)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10853..10872
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10932..10948)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10932..10948)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10945..10967)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10956..10972
                     /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(10980..10986,10987..11004,11005..11010))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11025..11046
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11163..11180)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11334..11922)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11414..11433)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12093..12884,12885..12953))
                     /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     12716..12735
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12954..13058)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(13133..13152)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(13324..13663,1..40)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
       61 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
      121 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
      181 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
      241 cgcgttttgc ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
      301 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
      361 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
      421 tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga
      481 ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg
      541 gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg
      601 cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag
      661 gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga
      721 acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg
      781 acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt
      841 agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga
      901 caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct
      961 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1021 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1081 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1141 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1201 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1261 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1321 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     1381 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     1441 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     1501 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     1561 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     1621 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     1681 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     1741 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     1801 tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt attcatccac
     1861 aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     1921 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt
     1981 cgggtttatt acagggacag cagagatcca gtttggttaa ttaaggtacc gagggcctat
     2041 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattagaa
     2101 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     2161 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     2221 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg
     2281 gagacgcttt tttcgtctca gtttgagagc tagaaatagc aagttcaaat aaggctagtc
     2341 cgttatcaac ttgaaaaagt ggcaccgagt cggtgcgctc acctattagc ggctaaggtt
     2401 ttttgaattc gctagctagg tcttgaaagg agtgggaatt ggctccggtg cccgtcagtg
     2461 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgatc
     2521 cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
     2581 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
     2641 ttttcgcaac gggtttgccg ccagaacaca ggaccggtgc caccatgaaa cggacagccg
     2701 acggaagcga gttcgagtca ccaaagaaga agcggaaagt ctctgaggtg gagttttccc
     2761 acgagtactg gatgagacat gccctgaccc tggccaagag ggcacgggat gagagggagg
     2821 tgcctgtggg agccgtgctg gtgctgaaca atagagtgat cggcgagggc tggaacagag
     2881 ccatcggcct gcacgaccca acagcccatg ccgaaattat ggccctgaga cagggcggcc
     2941 tggtcatgca gaactacaga ctgattgacg ccaccctgta cgtgacattc gagccttgcg
     3001 tgatgtgcgc cggcgccatg atccactcta ggatcggccg cgtggtgttt ggatggagaa
     3061 attctaaaag aggcgccgca ggctccctga tgaacgtgct gaactacccc ggcatgaatc
     3121 accgcgtcga aattaccgag ggaatcctgg cagatgaatg tgccgccctg ctgtgcgatt
     3181 tctatcggat gcctagacag gtgttcaatg ctcagaagaa ggcccagagc tccatcaact
     3241 ctggaggatc tagcggagga tcctctggca gcgagacacc aggaacaagc gagtcagcaa
     3301 caccagagag cagtggcggc agcagcggcg gcagcgacaa gaagtacagc atcggcctgg
     3361 ccatcggcac caactctgtg ggctgggccg tgatcaccga cgagtacaag gtgcccagca
     3421 agaaattcaa ggtgctgggc aacaccgacc ggcacagcat caagaagaac ctgatcggag
     3481 ccctgctgtt cgacagcggc gaaacagccg aggccacccg gctgaagaga accgccagaa
     3541 gaagatacac cagacggaag aaccggatct gctatctgca agagatcttc agcaacgaga
     3601 tggccaaggt ggacgacagc ttcttccaca gactggaaga gtccttcctg gtggaagagg
     3661 ataagaagca cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg
     3721 agaagtaccc caccatctac cacctgagaa agaaactggt ggacagcacc gacaaggccg
     3781 acctgcggct gatctatctg gccctggccc acatgatcaa gttccggggc cacttcctga
     3841 tcgagggcga cctgaacccc gacaacagcg acgtggacaa gctgttcatc cagctggtgc
     3901 agacctacaa ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg
     3961 ccatcctgtc tgccagactg agcaagagca gacggctgga aaatctgatc gcccagctgc
     4021 ccggcgagaa gaagaatggc ctgttcggaa acctgattgc cctgagcctg ggcctgaccc
     4081 ccaacttcaa gagcaacttc gacctggccg aggatgccaa actgcagctg agcaaggaca
     4141 cctacgacga cgacctggac aacctgctgg cccagatcgg cgaccagtac gccgacctgt
     4201 ttctggccgc caagaacctg tccgacgcca tcctgctgag cgacatcctg agagtgaaca
     4261 ccgagatcac caaggccccc ctgagcgcct ctatgatcaa gagatacgac gagcaccacc
     4321 aggacctgac cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag tacaaagaga
     4381 ttttcttcga ccagagcaag aacggctacg ccggctacat tgacggcgga gccagccagg
     4441 aagagttcta caagttcatc aagcccatcc tggaaaagat ggacggcacc gaggaactgc
     4501 tcgtgaagct gaacagagag gacctgctgc ggaagcagcg gaccttcgac aacggcagca
     4561 tcccccacca gatccacctg ggagagctgc acgccattct gcggcggcag gaagattttt
     4621 acccattcct gaaggacaac cgggaaaaga tcgagaagat cctgaccttc cgcatcccct
     4681 actacgtggg ccctctggcc aggggaaaca gcagattcgc ctggatgacc agaaagagcg
     4741 aggaaaccat caccccctgg aacttcgagg aagtggtgga caagggcgct tccgcccaga
     4801 gcttcatcga gcggatgacc aacttcgata agaacctgcc caacgagaag gtgctgccca
     4861 agcacagcct gctgtacgag tacttcaccg tgtataacga gctgaccaaa gtgaaatacg
     4921 tgaccgaggg aatgagaaag cccgccttcc tgagcggcga gcagaaaaag gccatcgtgg
     4981 acctgctgtt caagaccaac cggaaagtga ccgtgaagca gctgaaagag gactacttca
     5041 agaaaatcga gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct
     5101 ccctgggcac ataccacgat ctgctgaaaa ttatcaagga caaggacttc ctggacaatg
     5161 aggaaaacga ggacattctg gaagatatcg tgctgaccct gacactgttt gaggacagag
     5221 agatgatcga ggaacggctg aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc
     5281 agctgaagcg gcggagatac accggctggg gcaggctgag ccggaagctg atcaacggca
     5341 tccgggacaa gcagtccggc aagacaatcc tggatttcct gaagtccgac ggcttcgcca
     5401 acagaaactt catgcagctg atccacgacg acagcctgac ctttaaagag gacatccaga
     5461 aagcccaggt gtccggccag ggcgatagcc tgcacgagca cattgccaat ctggccggca
     5521 gccccgccat taagaagggc atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag
     5581 tgatgggccg gcacaagccc gagaacatcg tgatcgaaat ggccagagag aaccagacca
     5641 cccagaaggg acagaagaac agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag
     5701 agctgggcag ccagatcctg aaagaacacc ccgtggaaaa cacccagctg cagaacgaga
     5761 agctgtacct gtactacctg cagaatgggc gggatatgta cgtggaccag gaactggaca
     5821 tcaaccggct gtccgactac gatgtggacc atatcgtgcc tcagagcttt ctgaaggacg
     5881 actccatcga caacaaggtg ctgaccagaa gcgacaagaa ccggggcaag agcgacaacg
     5941 tgccctccga agaggtcgtg aagaagatga agaactactg gcggcagctg ctgaacgcca
     6001 agctgattac ccagagaaag ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg
     6061 aactggataa ggccggcttc atcaagagac agctggtgga aacccggcag atcacaaagc
     6121 acgtggcaca gatcctggac tcccggatga acactaagta cgacgagaat gacaagctga
     6181 tccgggaagt gaaagtgatc accctgaagt ccaagctggt gtccgatttc cggaaggatt
     6241 tccagtttta caaagtgcgc gagatcaaca actaccacca cgcccacgac gcctacctga
     6301 acgccgtcgt gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt
     6361 acggcgacta caaggtgtac gacgtgcgga agatgatcgc caagagcgag caggaaatcg
     6421 gcaaggctac cgccaagtac ttcttctaca gcaacatcat gaactttttc aagaccgaga
     6481 ttaccctggc caacggcgag atccggaagc ggcctctgat cgagacaaac ggcgaaaccg
     6541 gggagatcgt gtgggataag ggccgggatt ttgccaccgt gcggaaagtg ctgagcatgc
     6601 cccaagtgaa tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc aaagagtcta
     6661 tcctgcccaa gaggaacagc gataagctga tcgccagaaa gaaggactgg gaccctaaga
     6721 agtacggcgg cttcctgtgg cccaccgtgg cctattctgt gctggtggtg gccaaagtgg
     6781 aaaagggcaa gtccaagaaa ctgaagagtg tgaaagagct gctggggatc accatcatgg
     6841 aaagaagcag cttcgagaag aatcccatcg actttctgga agccaagggc tacaaagaag
     6901 tgaaaaagga cctgatcatc aagctgccta agtactccct gttcgagctg gaaaacggcc
     6961 ggaagagaat gctggcctct gccaagcagc tgcagaaggg aaacgaactg gccctgccct
     7021 ccaaatatgt gaacttcctg tacctggcca gccactatga gaagctgaag ggctcccccg
     7081 aggataatga gcagaaacag ctgtttgtgg aacagcacaa gcactacctg gacgagatca
     7141 tcgagcagat cagcgagttc tccaagagag tgatcctggc cgacgctaat ctggacaaag
     7201 tgctgtccgc ctacaacaag caccgggata agcccatcag agagcaggcc gagaatatca
     7261 tccacctgtt taccctgacc aatctgggag cccctgccgc cttcaagtac tttgacacca
     7321 ccatcgaccg gaagcagtac agaagcacca aagaggtgct ggacgccacc ctgatccacc
     7381 agagcatcac cggcctgtac gagacacgga tcgacctgtc tcagctggga ggtgacagcg
     7441 gaggctccaa aagaaccgcc gacggcagcg agttcgagcc caagaagaag aggaaagtca
     7501 gcgctggcgc aacaaacttc tctctgctga aacaagccgg agatgtcgaa gagaatcctg
     7561 gaccgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg
     7621 tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg
     7681 accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg
     7741 acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg
     7801 agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg
     7861 gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg
     7921 agcccgcgtg gttcctggcc accgtcggag tctcgcccga ccaccagggc aagggtctgg
     7981 gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc
     8041 tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg
     8101 ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct
     8161 gaacgcgtta agtcgacaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     8221 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     8281 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     8341 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     8401 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     8461 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     8521 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     8581 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     8641 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     8701 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     8761 ccccgcgtcg actttaagac caatgactta caaggcagct gtagatctta gccacttttt
     8821 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag atctgctttt
     8881 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     8941 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     9001 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     9061 aatctctagc agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     9121 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     9181 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     9241 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     9301 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     9361 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     9421 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     9481 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     9541 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     9601 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     9661 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     9721 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     9781 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     9841 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     9901 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     9961 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
    10021 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
    10081 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
    10141 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
    10201 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
    10261 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
    10321 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
    10381 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
    10441 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
    10501 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
    10561 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
    10621 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
    10681 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
    10741 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
    10801 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
    10861 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
    10921 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
    10981 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
    11041 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
    11101 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
    11161 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
    11221 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
    11281 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
    11341 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
    11401 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
    11461 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
    11521 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
    11581 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
    11641 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
    11701 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
    11761 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
    11821 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
    11881 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
    11941 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
    12001 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
    12061 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
    12121 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
    12181 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
    12241 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
    12301 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
    12361 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
    12421 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
    12481 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
    12541 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
    12601 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
    12661 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
    12721 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
    12781 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
    12841 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
    12901 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
    12961 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
    13021 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
    13081 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
    13141 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
    13201 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
    13261 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
    13321 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
    13381 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
    13441 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
    13501 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
    13561 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
    13621 cctggcatta tgcccagtac atgaccttat gggactttcc tac
//
