LOCUS       sequence-342241-       13010 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing Cas9 and a sgRNA against the human RB1 
            gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13010)
  AUTHORS   Chaikovsky AC, Li C, Jeng EE, Loebell S, Lee MC, Murray CW, Cheng R,
            Demeter J, Swaney DL, Chen SH, Newton BW, Johnson JR, Drainas AP, 
            Shue YT, Seoane JA, Srinivasan P, He A, Yoshida A, Hipkins SQ, 
            McCrea E, Poltorack CD, Krogan NJ, Diehl JA, Kong C, Jackson PK, 
            Curtis C, Petrov DA, Bassik MC, Winslow MM, Sage J
  TITLE     The AMBRA1 E3 ligase adaptor regulates the stability of cyclin D.
  JOURNAL   Nature. 2021 Apr;592(7856):794-798. doi: 10.1038/s41586-021-03474-7.
            Epub 2021 Apr 14.
  PUBMED    33854239
REFERENCE   2  (bases 1 to 13010)
  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: 
            "10.1038/s41586-021-03474-7"; journalName: "Nature"; date: 
            "2021-04"; volume: "592"; issue: "7856"; pages: "794-798"
COMMENT     Sequence Label: lentiCRISPRv2_RB1#1
FEATURES             Location/Qualifiers
     source          1..13010
                     /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        2608..2856
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2608..2628
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2779..2798
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2875..2950
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3012..3223
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3242..3251
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3248..7351
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3248..7351
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7352..7399
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7400..7423
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7433..7489
                     /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             7490..8086
                     /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     7983..8003
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8102..8690
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8155..8175)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8573..8584)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8762..8995
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9021..9038)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9027..9251
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9297..9725
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9384..9403)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9594..9615
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9734..9754)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9739..10068
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9919..10054
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9981..10000
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10116..10163
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10182..10556
                     /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    10686..10807
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10723..10742)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10777..10796
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10856..10872)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10856..10872)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10869..10891)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10880..10896
                     /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(10904..10910,10911..10928,10929..10934))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10949..10970
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11087..11104)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11258..11846)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11338..11357)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12017..12808,12809..12877))
                     /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     12640..12659
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12878..12982)
                     /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 aggtaccgag ggcctatttc ccatgattcc ttcatatttg
     2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag
     2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2821 ttggctttat atatcttgtg gaaaggacga aacaccggga gaaagtttca tctggtttta
     2881 gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc
     2941 gagtcggtgc ttttttgaat tcgctagcta ggtcttgaaa ggagtgggaa ttggctccgg
     3001 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
     3061 cggcaattga tccggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
     3121 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
     3181 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggaccggt tctagagcgc
     3241 tgccaccatg gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg
     3301 ggccgtgatc accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac
     3361 cgaccggcac agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac
     3421 agccgaggcc acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg
     3481 gatctgctat ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt
     3541 ccacagactg gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat
     3601 cttcggcaac atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct
     3661 gagaaagaaa ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct
     3721 ggcccacatg atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa
     3781 cagcgacgtg gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga
     3841 aaaccccatc aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa
     3901 gagcagacgg ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt
     3961 cggaaacctg attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct
     4021 ggccgaggat gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct
     4081 gctggcccag atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga
     4141 cgccatcctg ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag
     4201 cgcctctatg atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct
     4261 cgtgcggcag cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg
     4321 ctacgccggc tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc
     4381 catcctggaa aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct
     4441 gctgcggaag cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga
     4501 gctgcacgcc attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga
     4561 aaagatcgag aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg
     4621 aaacagcaga ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt
     4681 cgaggaagtg gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt
     4741 cgataagaac ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt
     4801 caccgtgtat aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc
     4861 cttcctgagc ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa
     4921 agtgaccgtg aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt
     4981 ggaaatctcc ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct
     5041 gaaaattatc aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga
     5101 tatcgtgctg accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac
     5161 ctatgcccac ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg
     5221 ctggggcagg ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac
     5281 aatcctggat ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca
     5341 cgacgacagc ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga
     5401 tagcctgcac gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct
     5461 gcagacagtg aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa
     5521 catcgtgatc gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg
     5581 cgagagaatg aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga
     5641 acaccccgtg gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa
     5701 tgggcgggat atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt
     5761 ggaccatatc gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac
     5821 cagaagcgac aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa
     5881 gatgaagaac tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga
     5941 caatctgacc aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa
     6001 gagacagctg gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg
     6061 gatgaacact aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct
     6121 gaagtccaag ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat
     6181 caacaactac caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat
     6241 caaaaagtac cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt
     6301 gcggaagatg atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt
     6361 ctacagcaac atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg
     6421 gaagcggcct ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg
     6481 ggattttgcc accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac
     6541 cgaggtgcag acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa
     6601 gctgatcgcc agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac
     6661 cgtggcctat tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa
     6721 gagtgtgaaa gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc
     6781 catcgacttt ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct
     6841 gcctaagtac tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg
     6901 cgaactgcag aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct
     6961 ggccagccac tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt
     7021 tgtggaacag cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa
     7081 gagagtgatc ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg
     7141 ggataagccc atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct
     7201 gggagcccct gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag
     7261 caccaaagag gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac
     7321 acggatcgac ctgtctcagc tgggaggcga caagcgacct gccgccacaa agaaggctgg
     7381 acaggctaag aagaagaaag attacaaaga cgatgacgat aagggatccg gcgcaacaaa
     7441 cttctctctg ctgaaacaag ccggagatgt cgaagagaat cctggaccga ccgagtacaa
     7501 gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc
     7561 cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg
     7621 ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg
     7681 ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg
     7741 ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc
     7801 gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct
     7861 ggccaccgtc ggagtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct
     7921 ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc
     7981 ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc
     8041 cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgaacgc gttaagtcga
     8101 caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     8161 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     8221 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     8281 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     8341 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     8401 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     8461 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     8521 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     8581 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     8641 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc gtcgacttta
     8701 agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga
     8761 ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct
     8821 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     8881 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     8941 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagggcc
     9001 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     9061 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     9121 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     9181 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     9241 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     9301 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     9361 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     9421 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     9481 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     9541 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     9601 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     9661 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     9721 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     9781 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     9841 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     9901 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     9961 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
    10021 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
    10081 gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt
    10141 atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg
    10201 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
    10261 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
    10321 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
    10381 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
    10441 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
    10501 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
    10561 tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
    10621 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
    10681 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
    10741 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
    10801 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
    10861 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
    10921 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
    10981 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
    11041 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
    11101 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
    11161 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
    11221 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
    11281 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
    11341 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
    11401 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
    11461 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
    11521 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
    11581 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
    11641 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
    11701 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
    11761 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
    11821 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
    11881 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
    11941 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
    12001 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
    12061 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
    12121 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
    12181 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
    12241 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
    12301 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
    12361 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
    12421 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
    12481 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
    12541 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
    12601 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
    12661 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
    12721 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
    12781 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
    12841 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
    12901 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
    12961 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
//
