lentiSAM v2 (Puro)

LOCUS       lentiSAM v2 (Puro)               13713 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  Modified version of lentiSAM v2, a lenti sgRNA cloning backbone with
            MS2 loops at tetraloop/stemloop 2, dCas9-VP64, and puro resistance 
            marker. Contains BsmBI sites for insertion of spacer sequences..
ACCESSION   .
VERSION     .
KEYWORDS    lentiSAM v2 (Puro)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13713)
  TITLE     Lenti CRISPR Activate
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 13713)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..13713
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(84..104)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        89..418
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      269..404
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     331..350
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        466..513
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             532..906
                     /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    1036..1157
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1073..1092)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1127..1146
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1206..1222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1206..1222)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1219..1241)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1230..1246
                     /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(1254..1284)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1299..1320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1437..1454)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1608..2196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1688..1707)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2367..3227)
                     /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     2990..3009
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3228..3332)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3407..3426)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3598..3977
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3979..4177
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4132..4152
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             4195..4375
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4422..4547
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5040..5273
                     /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    5800..5917
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5967..6207
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     5967..5987
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6138..6157
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        6253..6271
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        6323..6341
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        6428..6639
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             6656..6703
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6704..10804
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             10859..10879
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10904..11053
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             11075..11131
                     /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             11132..11728
                     /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     11625..11645
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    11765..12353
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(11818..11838)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(12236..12247)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(12356..12372)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12357..12373)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             12878..13058
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(13084..13101)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    13090..13314
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      13360..75
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(13447..13466)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     13657..13678
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
       61 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
      121 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
      181 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
      241 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
      301 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
      361 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
      421 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
      481 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag
      541 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
      601 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
      661 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
      721 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
      781 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
      841 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
      901 gactgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
      961 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     1021 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     1081 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     1141 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     1201 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     1261 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     1321 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     1381 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     1441 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     1501 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     1561 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     1621 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     1681 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     1741 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     1801 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     1861 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1921 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     1981 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     2041 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     2101 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     2161 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     2221 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     2281 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     2341 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     2401 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     2461 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     2521 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     2581 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     2641 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     2701 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     2761 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     2821 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     2881 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     2941 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     3001 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     3061 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     3121 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     3181 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     3241 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     3301 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     3361 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
     3421 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     3481 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
     3541 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
     3601 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
     3661 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     3721 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     3781 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     3841 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     3901 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     3961 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
     4021 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
     4081 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
     4141 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
     4201 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     4261 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     4321 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     4381 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     4441 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     4501 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     4561 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     4621 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     4681 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     4741 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     4801 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     4861 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     4921 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     4981 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     5041 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     5101 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     5161 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     5221 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     5281 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     5341 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     5401 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     5461 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     5521 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     5581 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     5641 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     5701 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     5761 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     5821 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     5881 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     5941 agatccagtt tggttaatta gctagcgagg gcctatttcc catgattcct tcatatttgc
     6001 atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta aacacaaaga
     6061 tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa
     6121 aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct
     6181 tggctttata tatcttgtgg aaaggacgaa acaccggaga cgggataccg tctctgtttt
     6241 agagctaggc caacatgagg atcacccatg tctgcagggc ctagcaagtt aaaataaggc
     6301 tagtccgtta tcaacttggc caacatgagg atcacccatg tctgcagggc caagtggcac
     6361 cgagtcggtg ctttttttgg atcctaggtc ttgaaaggag tgggaattgg ctccggtgcc
     6421 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
     6481 aattgatccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
     6541 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
     6601 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg cgtacggcca ccatgaaaag
     6661 gccggcggcc acgaaaaagg ccggccaggc aaaaaagaaa aaggacaaga agtacagcat
     6721 cggcctggcc atcggcacca actctgtggg ctgggccgtg atcaccgacg agtacaaggt
     6781 gcccagcaag aaattcaagg tgctgggcaa caccgaccgg cacagcatca agaagaacct
     6841 gatcggagcc ctgctgttcg acagcggcga aacagccgag gccacccggc tgaagagaac
     6901 cgccagaaga agatacacca gacggaagaa ccggatctgc tatctgcaag agatcttcag
     6961 caacgagatg gccaaggtgg acgacagctt cttccacaga ctggaagagt ccttcctggt
     7021 ggaagaggat aagaagcacg agcggcaccc catcttcggc aacatcgtgg acgaggtggc
     7081 ctaccacgag aagtacccca ccatctacca cctgagaaag aaactggtgg acagcaccga
     7141 caaggccgac ctgcggctga tctatctggc cctggcccac atgatcaagt tccggggcca
     7201 cttcctgatc gagggcgacc tgaaccccga caacagcgac gtggacaagc tgttcatcca
     7261 gctggtgcag acctacaacc agctgttcga ggaaaacccc atcaacgcca gcggcgtgga
     7321 cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc
     7381 ccagctgccc ggcgagaaga agaatggcct gttcggcaac ctgattgccc tgagcctggg
     7441 cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag
     7501 caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc
     7561 cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag
     7621 agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga
     7681 gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta
     7741 caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc
     7801 cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga
     7861 ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa
     7921 cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga
     7981 agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg
     8041 catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag
     8101 aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc
     8161 cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt
     8221 gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt
     8281 gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc
     8341 catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga
     8401 ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt
     8461 caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct
     8521 ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga
     8581 ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt
     8641 gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat
     8701 caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg
     8761 cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga
     8821 catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct
     8881 ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct
     8941 cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa
     9001 ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg
     9061 catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca
     9121 gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga
     9181 actggacatc aaccggctgt ccgactacga tgtggaccac atcgtgcctc agagctttct
     9241 gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaaggccc ggggcaagag
     9301 cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct
     9361 gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg
     9421 cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat
     9481 cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga
     9541 caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg
     9601 gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc
     9661 ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga
     9721 gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca
     9781 ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa
     9841 gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg
     9901 cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct
     9961 gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa
    10021 agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga
    10081 ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc
    10141 caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc tggggatcac
    10201 catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta
    10261 caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga
    10321 aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc
    10381 cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg
    10441 ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga
    10501 cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct
    10561 ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga
    10621 gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt
    10681 tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct
    10741 gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg
    10801 cgacagcgct ggaggaggtg gaagcggagg aggaggaagc ggaggaggag gtagcggacc
    10861 taagaaaaag aggaaggtgg cggccgctgg atccggacgg gctgacgcat tggacgattt
    10921 tgatctggat atgctgggaa gtgacgccct cgatgatttt gaccttgaca tgcttggttc
    10981 ggatgccctt gatgactttg acctcgacat gctcggcagt gacgcccttg atgatttcga
    11041 cctggacatg ctgattaact gtacaggcag tggagcaaca aacttctctc tgctgaaaca
    11101 agccggagat gtcgaagaga atcctggacc gaccgagtac aagcccacgg tgcgcctcgc
    11161 cacccgcgac gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc
    11221 cgccacgcgc cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga
    11281 actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc
    11341 cgcggtggcg gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat
    11401 cggcccgcgc atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaagg
    11461 cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggagtctc
    11521 gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggcggc
    11581 cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc tccccttcta
    11641 cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg
    11701 gtgcatgacc cgcaagcccg gtgcctgaac gcgttaagtc gaattcgata tcaagcttat
    11761 cggtaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
    11821 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
    11881 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
    11941 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
    12001 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
    12061 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
    12121 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
    12181 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
    12241 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
    12301 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata
    12361 ccgtcgacct cgagacctag aaaaacatgg agcaatcaca agtagcaata cagcagctac
    12421 caatgctgat tgtgcctggc tagaagcaca agaggaggag gaggtgggtt ttccagtcac
    12481 acctcaggta cctttaagac caatgactta caaggcagct gtagatctta gccacttttt
    12541 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag atatccttga
    12601 tctgtggatc taccacacac aaggctactt ccctgattgg cagaactaca caccagggcc
    12661 agggatcaga tatccactga cctttggatg gtgctacaag ctagtaccag ttgagcaaga
    12721 gaaggtagaa gaagccaatg aaggagagaa cacccgcttg ttacaccctg tgagcctgca
    12781 tgggatggat gacccggaga gagaagtatt agagtggagg tttgacagcc gcctagcatt
    12841 tcatcacatg gcccgagagc tgcatccgga ctgtactggg tctctctggt tagaccagat
    12901 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
    12961 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
    13021 cctcagaccc ttttagtcag tgtggaaaat ctctagcagg gcccgtttaa acccgctgat
    13081 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
    13141 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
    13201 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
    13261 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg
    13321 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat
    13381 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
    13441 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
    13501 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
    13561 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
    13621 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
    13681 caacactcaa ccctatctcg gtctattctt ttg
//