LentiCRISPRv2-mCherry

LOCUS        LentiCRISPRv2-mCherry               14984 bp ds-DNA     circular SYN 19-JAN-2022
DEFINITION  Lentiviral vector encoding sgRNA cloning site + 
            hSpCAS9-P2A-mCherry..
ACCESSION   .
VERSION     .
KEYWORDS    LentiCRISPRv2-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14984)
  TITLE     LentiCRISPRv2-mCherry
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14984)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 19, 2022 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..14984
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(2..21)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     248..268
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(277..705)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(387..408)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     599..618
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    751..975
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     964..981
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             1007..1240
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     misc_feature    1312..1900
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1418..1429
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1827..1847
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1916..2623)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2194..2213)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2463..2481
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2624..2680)
                     /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             complement(2690..2713)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2714..2761)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(2762..6865)
                     /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"
     regulatory      6862..6871
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     promoter        complement(6890..7101)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_RNA        complement(7163..7238)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(9128..9368)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     complement(9178..9197)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(9348..9368)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    9419..9536
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(9834..9878)
                     /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    10063..10296
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    10789..10914
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             10961..11141
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(11159..11357)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(11184..11204)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        11359..11738
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     11910..11929
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        12004..12108
                     /gene="bla"
                     /label=AmpR promoter
     CDS             12109..12969
                     /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     complement(12327..12346)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      13140..13728
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     13629..13648
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     13882..13899
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    14016..14037
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        14052..14082
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    14090..14106
                     /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)."
     primer_bind     14095..14117
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     14114..14130
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     14114..14130
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    14179..14300
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(14190..14209)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     14244..14263
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(14430..14804)
                     /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"
     promoter        complement(14823..14870)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(14918..263)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      14932..83
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa
       61 ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa
      121 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg
      181 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      241 tttccacacc ctaactgaca cacattccac agaattaatt cgcgttaaat ttttgttaaa
      301 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
      361 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
      421 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
      481 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
      541 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
      601 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
      661 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtgggga taccccctag
      721 agccccagct ggttctttcc gcctcagaag ccatagagcc caccgcatcc ccagcatgcc
      781 tgctattgtc ttcccaatcc tcccccttgc tgtcctgccc caccccaccc cccagaatag
      841 aatgacacct actcagacaa tgcgatgcaa tttcctcatt ttattaggaa aggacagtgg
      901 gagtggcacc ttccagggtc aaggaaggca cgggggaggg gcaaacaaca gatggctggc
      961 aactagaagg cacagtcgag gctgatcagc gggtttaaac gggccctgct agagattttc
     1021 cacactgact aaaagggtct gagggatctc tagttaccag agtcacacaa cagacgggca
     1081 cacactactt gaagcactca aggcaagctt tattgaggct taagcagtgg gttccctagt
     1141 tagccagaga gctcccaggc tcagatctgg tctaaccaga gagacccagt acaagcaaaa
     1201 agcagatctt gtcttcgttg ggagtgaatt agcccttcca gtcccccctt ttcttttaaa
     1261 aagtggctaa gatctacagc tgccttgtaa gtcattggtc ttaaagtcga cgcggggagg
     1321 cggcccaaag ggagatccga ctcgtctgag ggcgaaggcg aagacgcgga agaggccgca
     1381 gagccggcag caggccgcgg gaaggaaggt ccgctggatt gagggccgaa gggacgtagc
     1441 agaaggacgt cccgcgcaga atccaggtgg caacacaggc gagcagccaa ggaaaggacg
     1501 atgatttccc cgacaacacc acggaattgt cagtgcccaa cagccgagcc cctgtccagc
     1561 agcgggcaag gcaggcggcg atgagttccg ccgtggcaat agggaggggg aaagcgaaag
     1621 tcccggaaag gagctgacag gtggtggcaa tgccccaacc agtgggggtt gcgtcagcaa
     1681 acacagtgca caccacgcca cgttgcctga caacgggcca caactcctca taaagagaca
     1741 gcaaccagga tttatacaag gaggagaaaa tgaaagccat acgggaagca atagcatgat
     1801 acaaaggcat taaagcagcg tatccacata gcgtaaaagg agcaacatag ttaagaatac
     1861 cagtcaatct ttcacaaatt ttgtaatcca gaggttgatt gtcgacttaa cgcgtttact
     1921 tgtacagctc gtccatgccg ccggtggagt ggcggccctc ggcgcgttcg tactgttcca
     1981 cgatggtgta gtcctcgttg tgggaggtga tgtccaactt gatgttgacg ttgtaggcgc
     2041 cgggcagctg cacgggcttc ttggccttgt aggtggtctt gacctcagcg tcgtagtggc
     2101 cgccgtcctt cagcttcagc ctctgcttga tctcgccctt cagggcgccg tcctcggggt
     2161 acatccgctc ggaggaggcc tcccagccca tggtcttctt ctgcattacg gggccgtcgg
     2221 aggggaagtt ggtgccgcgc agcttcacct tgtagatgaa ctcgccgtcc tgcagggagg
     2281 agtcctgggt cacggtcacc acgccgccgt cctcgaagtt catcacgcgc tcccacttga
     2341 agccctcggg gaaggacagc ttcaagtagt cggggatgtc ggcggggtgc ttcacgtagg
     2401 ccttggagcc gtacatgaac tgaggggaca ggatgtccca ggcgaagggc agggggccac
     2461 ccttggtcac cttcagcttg gcggtctggg tgccctcgta ggggcggccc tcgccctcgc
     2521 cctcgatctc gaactcgtgg ccgttcacgg agccctccat gtgcaccttg aagcgcatga
     2581 actccttgat gatggccatg ttatcctcct cgcccttgct caccggtcca ggattctctt
     2641 cgacatctcc ggcttgtttc agcagagaga agtttgttgc gccggatccc ttatcgtcat
     2701 cgtctttgta atctttcttc ttcttagcct gtccagcctt ctttgtggcg gcaggtcgct
     2761 tgtcgcctcc cagctgagac aggtcgatcc gtgtctcgta caggccggtg atgctctggt
     2821 ggatcagggt ggcgtccagc acctctttgg tgctggtgta cctcttccgg tcgatggtgg
     2881 tgtcaaagta cttgaaggcg gcaggggctc ccagattggt cagggtaaac aggtggatga
     2941 tattctcggc ctgctctctg atgggcttat cccggtgctt gttgtaggcg gacagcactt
     3001 tgtccagatt agcgtcggcc aggatcactc tcttggagaa ctcgctgatc tgctcgatga
     3061 tctcgtccag gtagtgcttg tgctgttcca caaacagctg tttctgctca ttatcctcgg
     3121 gggagccctt cagcttctca tagtggctgg ccaggtacag gaagttcaca tatttggagg
     3181 gcagggccag ttcgtttccc ttctgcagtt cgccggcaga ggccagcatt ctcttccggc
     3241 cgttttccag ctcgaacagg gagtacttag gcagcttgat gatcaggtcc tttttcactt
     3301 ctttgtagcc cttggcttcc agaaagtcga tgggattctt ctcgaagctg cttctttcca
     3361 tgatggtgat ccccagcagc tctttcacac tcttcagttt cttggacttg cccttttcca
     3421 ctttggccac caccagcaca gaataggcca cggtggggct gtcgaagccg ccgtacttct
     3481 tagggtccca gtccttcttt ctggcgatca gcttatcgct gttcctcttg ggcaggatag
     3541 actctttgct gaagccgcct gtctgcacct cggtcttttt cacgatattc acttggggca
     3601 tgctcagcac tttccgcacg gtggcaaaat cccggccctt atcccacacg atctccccgg
     3661 tttcgccgtt tgtctcgatc agaggccgct tccggatctc gccgttggcc agggtaatct
     3721 cggtcttgaa aaagttcatg atgttgctgt agaagaagta cttggcggta gccttgccga
     3781 tttcctgctc gctcttggcg atcatcttcc gcacgtcgta caccttgtag tcgccgtaca
     3841 cgaactcgct ttccagctta gggtactttt tgatcagggc ggttcccacg acggcgttca
     3901 ggtaggcgtc gtgggcgtgg tggtagttgt tgatctcgcg cactttgtaa aactggaaat
     3961 ccttccggaa atcggacacc agcttggact tcagggtgat cactttcact tcccggatca
     4021 gcttgtcatt ctcgtcgtac ttagtgttca tccgggagtc caggatctgt gccacgtgct
     4081 ttgtgatctg ccgggtttcc accagctgtc tcttgatgaa gccggcctta tccagttcgc
     4141 tcaggccgcc tctctcggcc ttggtcagat tgtcgaactt tctctgggta atcagcttgg
     4201 cgttcagcag ctgccgccag tagttcttca tcttcttcac gacctcttcg gagggcacgt
     4261 tgtcgctctt gccccggttc ttgtcgcttc tggtcagcac cttgttgtcg atggagtcgt
     4321 ccttcagaaa gctctgaggc acgatatggt ccacatcgta gtcggacagc cggttgatgt
     4381 ccagttcctg gtccacgtac atatcccgcc cattctgcag gtagtacagg tacagcttct
     4441 cgttctgcag ctgggtgttt tccacggggt gttctttcag gatctggctg cccagctctt
     4501 tgatgccctc ttcgatccgc ttcattctct cgcggctgtt cttctgtccc ttctgggtgg
     4561 tctggttctc tctggccatt tcgatcacga tgttctcggg cttgtgccgg cccatcactt
     4621 tcacgagctc gtccaccacc ttcactgtct gcaggatgcc cttcttaatg gcggggctgc
     4681 cggccagatt ggcaatgtgc tcgtgcaggc tatcgccctg gccggacacc tgggctttct
     4741 ggatgtcctc tttaaaggtc aggctgtcgt cgtggatcag ctgcatgaag tttctgttgg
     4801 cgaagccgtc ggacttcagg aaatccagga ttgtcttgcc ggactgcttg tcccggatgc
     4861 cgttgatcag cttccggctc agcctgcccc agccggtgta tctccgccgc ttcagctgct
     4921 tcatcacttt gtcgtcgaac aggtgggcat aggttttcag ccgttcctcg atcatctctc
     4981 tgtcctcaaa cagtgtcagg gtcagcacga tatcttccag aatgtcctcg ttttcctcat
     5041 tgtccaggaa gtccttgtcc ttgataattt tcagcagatc gtggtatgtg cccagggagg
     5101 cgttgaaccg atcttccacg ccggagattt ccacggagtc gaagcactcg attttcttga
     5161 agtagtcctc tttcagctgc ttcacggtca ctttccggtt ggtcttgaac agcaggtcca
     5221 cgatggcctt tttctgctcg ccgctcagga aggcgggctt tctcattccc tcggtcacgt
     5281 atttcacttt ggtcagctcg ttatacacgg tgaagtactc gtacagcagg ctgtgcttgg
     5341 gcagcacctt ctcgttgggc aggttcttat cgaagttggt catccgctcg atgaagctct
     5401 gggcggaagc gcccttgtcc accacttcct cgaagttcca gggggtgatg gtttcctcgc
     5461 tctttctggt catccaggcg aatctgctgt ttcccctggc cagagggccc acgtagtagg
     5521 ggatgcggaa ggtcaggatc ttctcgatct tttcccggtt gtccttcagg aatgggtaaa
     5581 aatcttcctg ccgccgcaga atggcgtgca gctctcccag gtggatctgg tgggggatgc
     5641 tgccgttgtc gaaggtccgc tgcttccgca gcaggtcctc tctgttcagc ttcacgagca
     5701 gttcctcggt gccgtccatc ttttccagga tgggcttgat gaacttgtag aactcttcct
     5761 ggctggctcc gccgtcaatg tagccggcgt agccgttctt gctctggtcg aagaaaatct
     5821 ctttgtactt ctcaggcagc tgctgccgca cgagagcttt cagcagggtc aggtcctggt
     5881 ggtgctcgtc gtatctcttg atcatagagg cgctcagggg ggccttggtg atctcggtgt
     5941 tcactctcag gatgtcgctc agcaggatgg cgtcggacag gttcttggcg gccagaaaca
     6001 ggtcggcgta ctggtcgccg atctgggcca gcaggttgtc caggtcgtcg tcgtaggtgt
     6061 ccttgctcag ctgcagtttg gcatcctcgg ccaggtcgaa gttgctcttg aagttggggg
     6121 tcaggcccag gctcagggca atcaggtttc cgaacaggcc attcttcttc tcgccgggca
     6181 gctgggcgat cagattttcc agccgtctgc tcttgctcag tctggcagac aggatggcct
     6241 tggcgtccac gccgctggcg ttgatggggt tttcctcgaa cagctggttg taggtctgca
     6301 ccagctggat gaacagcttg tccacgtcgc tgttgtcggg gttcaggtcg ccctcgatca
     6361 ggaagtggcc ccggaacttg atcatgtggg ccagggccag atagatcagc cgcaggtcgg
     6421 ccttgtcggt gctgtccacc agtttctttc tcaggtggta gatggtgggg tacttctcgt
     6481 ggtaggccac ctcgtccacg atgttgccga agatggggtg ccgctcgtgc ttcttatcct
     6541 cttccaccag gaaggactct tccagtctgt ggaagaagct gtcgtccacc ttggccatct
     6601 cgttgctgaa gatctcttgc agatagcaga tccggttctt ccgtctggtg tatcttcttc
     6661 tggcggttct cttcagccgg gtggcctcgg ctgtttcgcc gctgtcgaac agcagggctc
     6721 cgatcaggtt cttcttgatg ctgtgccggt cggtgttgcc cagcaccttg aatttcttgc
     6781 tgggcacctt gtactcgtcg gtgatcacgg cccagcccac agagttggtg ccgatgtcca
     6841 ggccgatgct gtacttcttg tccatggtgg cagcgctcta gaaccggtcc tgtgttctgg
     6901 cggcaaaccc gttgcgaaaa agaacgttca cggcgactac tgcacttata tacggttctc
     6961 ccccaccctc gggaaaaagg cggagccagt acacgacatc actttcccag tttaccccgc
     7021 gccaccttct ctaggcaccg gatcaattgc cgacccctcc ccccaacttc tcggggactg
     7081 tgggcgatgt gcgctctgcc cactgacggg caccggagcc aattcccact cctttcaaga
     7141 cctagctagc gaattcaaaa aagcaccgac tcggtgccac tttttcaagt tgataacgga
     7201 ctagccttat tttaacttgc tatttctagc tctaaaacag agacgtacaa aaaagagcaa
     7261 gaagctaaaa aagatttaaa aattattttt agcgcagtta atggaacagg aactaaattt
     7321 accccaaaaa tattacgtga atcaggatat aacgttattg aggttgaaga gcatgcattt
     7381 gaagatgaaa catttaaaaa tgttgtaaat ccaaatccag aatttgatcc tgcatgaaaa
     7441 ataccgcttg aatatggtat taaacatgat gcagatatta ttattatgaa tgacccagat
     7501 gctgacagat ttggaatggc aataaaacat gatggtcatt ttgtaagatt agatggaaat
     7561 caaacaggac caattttaat tgattgaaaa ttatcaaatc taaaacgctt aaatagcatt
     7621 ccaaaaaatc cggctctata ttcaagtttt gtaacaagtg atttgggtga tagaatcgct
     7681 catgaaaaat atggagttaa tattgtaaaa actttaactg gatttaaatg aatgggtaga
     7741 gaaattgcta aagaagaaga taacggatta aattttgttt ttgcttatga agaaagttat
     7801 ggatatgtaa ttgatgactc agctagagat aaagatggaa tacaagcttc tatattaata
     7861 gcagaggctg cttgatttta taaaaaacaa aataaaacat tagtagacta tttagaagat
     7921 ttatttaaag aaatgggtgc atattacact ttcactttaa acttgaattt taaaccagaa
     7981 gaaaagaaat taaaaattga accattaatg aaatcattga gagcaacacc cttaactcaa
     8041 attgctggac ttaaagttgt taatgttgaa gactacatcg atggaatgta taatatgcca
     8101 ggacaagact tactaaaatt ttatttagaa gataagtcat gatttgctgt tcgcccaagt
     8161 ggaactgaac ctaaactaaa aatttatttt ataggtgttg gtgaatctgt tcaaaacgct
     8221 aaagttaaag tagacgaaat tattaaagaa ttaaaattaa aaatgaatat ataggagaaa
     8281 aaatgaaact aaacaaatat atagatcaca cattattaaa acaagatgct acgaaagctg
     8341 aaattaaaca attatgtgat gaagcaattg aatttgattt tgcaacagtt tgtgttaatt
     8401 catattgaac aagctattgt aaagaattat taaaaggcac aaatgtagga ataacaaatg
     8461 ttgtaggttt tcctctaggt gcatgcacaa cagctacaaa agcattcgaa gtttctgaag
     8521 caattaaaga tggtgcaaca gaaattgata tggtattaaa tattggtgca ttaaaagaca
     8581 aaaattatga attagtttta gaagacatga aagctgtaaa aaaagcagct ggatcacatg
     8641 ttgttaaatg tattatggaa aattgtttat taacaaaaga agaaatcatg aaagcttgtg
     8701 aaatagctgt tgaagctgga ttagaatttg ttaaaacatc aacaggattt tcaaaatcag
     8761 gtgcaacatt tgaagatgtt aaactaatga agtcagttgt taaagacaat gctttagtta
     8821 aagcagctgg tggagttaga acatttgaag atgctcaaaa aatgattgaa gcaggagctg
     8881 accgcttagg aacaagtggt ggagtagcta ttattaaagg tgaagaaaac aacgcgagtt
     8941 actaaaacta gcgttttttt attttgctca tttttattaa aagtttgcaa aaaggaacat
     9001 aaaaattcta attattgata ctaaagttat taaaaagaag attttggttg attttataaa
     9061 ggtcatagaa tataatattt tagcatgtgt attttgtgtg ctcatttaca accgtctccg
     9121 gtgtttcgtc ctttccacaa gatatataaa gccaagaaat cgaaatactt tcaagttacg
     9181 gtaagcatat gatagtccat tttaaaacat aattttaaaa ctgcaaacta cccaagaaat
     9241 tattactttc tacgtcacgt attttgtact aatatctttg tgtttacagt caaattaatt
     9301 ctaattatct ctctaacagc cttgtatcgt atatgcaaat atgaaggaat catgggaaat
     9361 aggccctcgg taccttaatt aaccaaactg gatctctgct gtccctgtaa taaacccgaa
     9421 aattttgaat ttttgtaatt tgtttttgta attctttagt ttgtatgtct gttgctatta
     9481 tgtctactat tctttcccct gcactgtacc ccccaatccc cccttttctt ttaaaattgt
     9541 ggatgaatac tgccatttgt ctgcagaatt ggcgcacgca gtgccgatcc gttcactaat
     9601 cgaatggatc tgtctctgtc tctctctcca ccttcttctt ctattccttc gggcctgtcg
     9661 ggtcccctcg gggttgggag gtgggtctga aacgataatg gtgaatatcc ctgcctaact
     9721 ctattcacta tagaaagtac agcaaaaact attcttaaac ctaccaagcc tcctactatc
     9781 attatgaata attttatata ccacagccaa tttgttatgt taaaccaatt ccacaaactt
     9841 gcccatttat ctaattccaa taattcttgt tcattctttt cttgctggtt ttgcgattct
     9901 tcaattaagg agtgtattaa gcttgtgtaa ttgttaattt ctctgtccca ctccatccag
     9961 gtcgtgtgat tccaaatctg ttccagagat ttattactcc aactagcatt ccaaggcaca
    10021 gcagtggtgc aaatgagttt tccagagcaa ccccaaatcc ccaggagctg ttgatccttt
    10081 aggtatcttt ccacagccag gattcttgcc tggagctgct tgatgcccca gactgtgagt
    10141 tgcaacagat gctgttgcgc ctcaatagcc ctcagcaaat tgttctgctg ctgcactata
    10201 ccagacaata attgtctggc ctgtaccgtc agcgtcattg acgctgcgcc catagtgctt
    10261 cctgctgctc ccaagaaccc aaggaacaaa gctcctattc ccactgctct tttttctctc
    10321 tgcaccactc ttctctttgc cttggtgggt gctactccta atggttcaat ttttactact
    10381 ttatatttat ataattcact tctccaattg tccctcatat ctcctcctcc aggtctgaag
    10441 atcagcggcc gcttgctgtg cggtggtctt acttttgttt tgctcttcct ctatcttgtc
    10501 taaagcttcc ttggtgtctt ttatctctat cctttgatgc acacaataga gggttgctac
    10561 tgtattatat aatgatctaa gttcttctga tcctgtctga agggatggtt gtagctgtcc
    10621 cagtatttgt ctacagcctt ctgatgtttc taacaggcca ggattaactg cgaatcgttc
    10681 tagctccctg cttgcccata ctatatgttt taatttatat tttttctttc cccctggcct
    10741 taaccgaatt ttttcccatc gcgatctaat tctcccccgc ttaatactga cgctctcgca
    10801 cccatctctc tccttctagc ctccgctagt caaaattttt ggcgtactca ccagtcgccg
    10861 cccctcgcct cttgccgtgc gcgcttcagc aagccgagtc ctgcgtcgag agagctcctc
    10921 tggtttccct ttcgctttca agtccctgtt cgggcgccac tgctagagat tttccacact
    10981 gactaaaagg gtctgaggga tctctagtta ccagagtcac acaacagacg ggcacacact
    11041 acttgaagca ctcaaggcaa gctttattga ggcttaagca gtgggttccc tagttagcca
    11101 gagagctccc aggctcagat ctggtctaac cagagagacc cagtacaggc aaaacgcgct
    11161 gcttatatag acctcccacc gtacacgcct accgcccatt tgcgtcaatg gggcggagtt
    11221 gttacgacat tttggaaagt cccgttgatt ttggtgccaa aacaaactcc cattgacgtc
    11281 aatggggtgg agacttggaa atccccgtga gtcaaaccgc tatccacgcc cattgatgta
    11341 ctgccaaaac cgcatcacca tggtaatagc gatgactaat acgtagatgt actgccaagt
    11401 aggaaagtcc cataaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt
    11461 gacgtcaata gggggcgtac ttggcatatg atacacttga tgtactgcca agtgggcagt
    11521 ttaccgtaaa tactccaccc attgacgtca atggaaagtc cctattggcg ttactatggg
    11581 aacatacgtc attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc
    11641 atttaccgta agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt
    11701 aattgattac tattaataac tagtcaataa tcaatgtcaa cgcgtatatc tggcccgtac
    11761 atcgcgaagc agcgcaaaac gcctaaccct aagcagattc ttcatgcaat tgtcggtcaa
    11821 gccttgcctt gttgtagctt aaattttgct cgcgcactac tcagcgacct ccaacacaca
    11881 agcagggagc agatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca
    11941 ccatagggga tcgggagatc tcccgatccg tcgacgtcag gtggcacttt tcggggaaat
    12001 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
    12061 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
    12121 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
    12181 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
    12241 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
    12301 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
    12361 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
    12421 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
    12481 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
    12541 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
    12601 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
    12661 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
    12721 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
    12781 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
    12841 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
    12901 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
    12961 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
    13021 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
    13081 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
    13141 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
    13201 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
    13261 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
    13321 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
    13381 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
    13441 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
    13501 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
    13561 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
    13621 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
    13681 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
    13741 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
    13801 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
    13861 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
    13921 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
    13981 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
    14041 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
    14101 taacaatttc acacaggaaa cagctatgac catgattacg ccaagctcta gctagaggtc
    14161 gacggtatac agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
    14221 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
    14281 taagctgcaa taaacaagtt ggggtgggcg aagaactcca gcatgagatc cccgcgctgg
    14341 aggatcatcc agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg
    14401 gcggtggaat cgaaatctcg tagcacgtgt cagtcctgct cctcggccac gaagtgcacg
    14461 cagttgccgg ccgggtcgcg cagggcgaac tcccgccccc acggctgctc gccgatctcg
    14521 gtcatggccg gcccggaggc gtcccggaag ttcgtggaca cgacctccga ccactcggcg
    14581 tacagctcgt ccaggccgcg cacccacacc caggccaggg tgttgtccgg caccacctgg
    14641 tcctggaccg cgctgatgaa cagggtcacg tcgtcccgga ccacaccggc gaagtcgtcc
    14701 tccacgaagt cccgggagaa cccgagccgg tcggtccaga actcgaccgc tccggcgacg
    14761 tcgcgcgcgg tgagcaccgg aacggcactg gtcaacttgg ccatggttta gttcctcacc
    14821 ttgtcgtatt atactatgcc gatatactat gccgatgatt aattgtcaac acgtgctgat
    14881 cagatccgaa aatggatata caagctcccg ggagcttttt gcaaaagcct aggcctccaa
    14941 aaaagcctcc tcactacttc tggaatagct cagaggcaga ggcg
//