LOCUS       sequence-384453-       11244 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses eCas9, BlastR and sgRNA cloning site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11244)
  AUTHORS   Freed-Pastor WA, Lambert LJ, Ely ZA, Pattada NB, Bhutkar A, Eng G, 
            Mercer KL, Garcia AP, Lin L, Rideout WM 3rd, Hwang WL, Schenkel JM, 
            Jaeger AM, Bronson RT, Westcott PMK, Hether TD, Divakar P, Reeves 
            JW, Deshpande V, Delorey T, Phillips D, Yilmaz OH, Regev A, Jacks T
  TITLE     The CD155/TIGIT axis promotes and maintains immune evasion in 
            neoantigen-expressing pancreatic cancer.
  JOURNAL   Cancer Cell. 2021 Oct 11;39(10):1342-1360.e14. doi: 
            10.1016/j.ccell.2021.07.007. Epub 2021 Aug 5.
  PUBMED    34358448
REFERENCE   2  (bases 1 to 11244)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.ccell.2021.07.007"; journalName: "Cancer Cell"; date: 
            "2021-10-11- 11"; volume: "39"; issue: "10"; pages: "1342-1360.e14"
COMMENT     Sequence Label: U6-sgfiller-eCas9-T2A-BlastR
FEATURES             Location/Qualifiers
     source          1..11244
                     /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    1684..1917
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2102..2146
                     /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    2444..2561
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2654..2902
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2654..2674
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2825..2844
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2946..3021
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3101..3312
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3344..3353
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3353..3418
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3425..3445
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(3470..6007,6008..6010,6011..6472,6473..6475,6476..6643,
                     6644..6646,6647..7570)
                     /codon_start=1
                     /product="Cas9 endonuclease from the Streptococcus pyogenes
                     Type II CRISPR/Cas system, mutated to improve targeting 
                     specificity (Slaymaker et al., 2016)"
                     /label=eSpCas9(1.1)
                     /note="carries the mutations K848A, K1003A, and R1060A"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPALESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKAPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7571..7618
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7628..7681
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             7682..8080
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8133..8721
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8186..8206)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8604..8615)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(8724..8740)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8725..8741)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9250..9430
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(9492..10080)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9572..9591)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(10251..11042,11043..11111))
                     /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     10874..10893
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11112..11216)
                     /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 gccgctgatc ttcagacctg gaggaggaga
     1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt
     1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg
     1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc
     1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa
     1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa
     1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg
     1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg
     1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga
     2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga
     2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa
     2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg
     2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc
     2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga
     2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact
     2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg
     2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     2581 gcagagatcc agtttggtta gtaccgggcc cgggtcgact agtgatcagt gtgagggagt
     2641 gtaaagctgg tttgagggcc tatttcccat gattccttca tatttgcata tacgatacaa
     2701 ggctgttaga gagataatta gaattaattt gactgtaaac acaaagatat tagtacaaaa
     2761 tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa
     2821 aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat
     2881 cttgtggaaa ggacgaaaca ccggagacgg ttgtaaatga gcacacaact tttttgtacg
     2941 tctctgtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa
     3001 aaagtggcac cgagtcggtg cttttttaaa ccgctgttcc taggaatccc gaggccttag
     3061 gtcttgaaag gagtgggaat tggctccggt gcccgtcagt gggcagagcg cacatcgccc
     3121 acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta gagaaggtgg
     3181 cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg
     3241 ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc
     3301 gccagaacac agggacccga cattagcgct acagcttaag cgggccacca tggactataa
     3361 ggaccacgac ggagactaca aggatcatga tattgattac aaagacgatg acgataagat
     3421 ggccccaaag aagaagcgga aggtcggtat ccacggagtc ccagcagccg acaagaagta
     3481 cagcatcggc ctggacatcg gcaccaactc tgtgggctgg gccgtgatca ccgacgagta
     3541 caaggtgccc agcaagaaat tcaaggtgct gggcaacacc gaccggcaca gcatcaagaa
     3601 gaacctgatc ggagccctgc tgttcgacag cggcgaaaca gccgaggcca cccggctgaa
     3661 gagaaccgcc agaagaagat acaccagacg gaagaaccgg atctgctatc tgcaagagat
     3721 cttcagcaac gagatggcca aggtggacga cagcttcttc cacagactgg aagagtcctt
     3781 cctggtggaa gaggataaga agcacgagcg gcaccccatc ttcggcaaca tcgtggacga
     3841 ggtggcctac cacgagaagt accccaccat ctaccacctg agaaagaaac tggtggacag
     3901 caccgacaag gccgacctgc ggctgatcta tctggccctg gcccacatga tcaagttccg
     3961 gggccacttc ctgatcgagg gcgacctgaa ccccgacaac agcgacgtgg acaagctgtt
     4021 catccagctg gtgcagacct acaaccagct gttcgaggaa aaccccatca acgccagcgg
     4081 cgtggacgcc aaggccatcc tgtctgccag actgagcaag agcagacggc tggaaaatct
     4141 gatcgcccag ctgcccggcg agaagaagaa tggcctgttc ggaaacctga ttgccctgag
     4201 cctgggcctg acccccaact tcaagagcaa cttcgacctg gccgaggatg ccaaactgca
     4261 gctgagcaag gacacctacg acgacgacct ggacaacctg ctggcccaga tcggcgacca
     4321 gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac gccatcctgc tgagcgacat
     4381 cctgagagtg aacaccgaga tcaccaaggc ccccctgagc gcctctatga tcaagagata
     4441 cgacgagcac caccaggacc tgaccctgct gaaagctctc gtgcggcagc agctgcctga
     4501 gaagtacaaa gagattttct tcgaccagag caagaacggc tacgccggct acattgacgg
     4561 cggagccagc caggaagagt tctacaagtt catcaagccc atcctggaaa agatggacgg
     4621 caccgaggaa ctgctcgtga agctgaacag agaggacctg ctgcggaagc agcggacctt
     4681 cgacaacggc agcatccccc accagatcca cctgggagag ctgcacgcca ttctgcggcg
     4741 gcaggaagat ttttacccat tcctgaagga caaccgggaa aagatcgaga agatcctgac
     4801 cttccgcatc ccctactacg tgggccctct ggccagggga aacagcagat tcgcctggat
     4861 gaccagaaag agcgaggaaa ccatcacccc ctggaacttc gaggaagtgg tggacaaggg
     4921 cgcttccgcc cagagcttca tcgagcggat gaccaacttc gataagaacc tgcccaacga
     4981 gaaggtgctg cccaagcaca gcctgctgta cgagtacttc accgtgtata acgagctgac
     5041 caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc ttcctgagcg gcgagcagaa
     5101 aaaggccatc gtggacctgc tgttcaagac caaccggaaa gtgaccgtga agcagctgaa
     5161 agaggactac ttcaagaaaa tcgagtgctt cgactccgtg gaaatctccg gcgtggaaga
     5221 tcggttcaac gcctccctgg gcacatacca cgatctgctg aaaattatca aggacaagga
     5281 cttcctggac aatgaggaaa acgaggacat tctggaagat atcgtgctga ccctgacact
     5341 gtttgaggac agagagatga tcgaggaacg gctgaaaacc tatgcccacc tgttcgacga
     5401 caaagtgatg aagcagctga agcggcggag atacaccggc tggggcaggc tgagccggaa
     5461 gctgatcaac ggcatccggg acaagcagtc cggcaagaca atcctggatt tcctgaagtc
     5521 cgacggcttc gccaacagaa acttcatgca gctgatccac gacgacagcc tgacctttaa
     5581 agaggacatc cagaaagccc aggtgtccgg ccagggcgat agcctgcacg agcacattgc
     5641 caatctggcc ggcagccccg ccattaagaa gggcatcctg cagacagtga aggtggtgga
     5701 cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac atcgtgatcg aaatggccag
     5761 agagaaccag accacccaga agggacagaa gaacagccgc gagagaatga agcggatcga
     5821 agagggcatc aaagagctgg gcagccagat cctgaaagaa caccccgtgg aaaacaccca
     5881 gctgcagaac gagaagctgt acctgtacta cctgcagaat gggcgggata tgtacgtgga
     5941 ccaggaactg gacatcaacc ggctgtccga ctacgatgtg gaccatatcg tgcctcagag
     6001 ctttctggcc gacgactcca tcgacaacaa ggtgctgacc agaagcgaca agaaccgggg
     6061 caagagcgac aacgtgccct ccgaagaggt cgtgaagaag atgaagaact actggcggca
     6121 gctgctgaac gccaagctga ttacccagag aaagttcgac aatctgacca aggccgagag
     6181 aggcggcctg agcgaactgg ataaggccgg cttcatcaag agacagctgg tggaaacccg
     6241 gcagatcaca aagcacgtgg cacagatcct ggactcccgg atgaacacta agtacgacga
     6301 gaatgacaag ctgatccggg aagtgaaagt gatcaccctg aagtccaagc tggtgtccga
     6361 tttccggaag gatttccagt tttacaaagt gcgcgagatc aacaactacc accacgccca
     6421 cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc aaaaagtacc ctgcgctgga
     6481 aagcgagttc gtgtacggcg actacaaggt gtacgacgtg cggaagatga tcgccaagag
     6541 cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc tacagcaaca tcatgaactt
     6601 tttcaagacc gagattaccc tggccaacgg cgagatccgg aaggcgcctc tgatcgagac
     6661 aaacggcgaa accggggaga tcgtgtggga taagggccgg gattttgcca ccgtgcggaa
     6721 agtgctgagc atgccccaag tgaatatcgt gaaaaagacc gaggtgcaga caggcggctt
     6781 cagcaaagag tctatcctgc ccaagaggaa cagcgataag ctgatcgcca gaaagaagga
     6841 ctgggaccct aagaagtacg gcggcttcga cagccccacc gtggcctatt ctgtgctggt
     6901 ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag agtgtgaaag agctgctggg
     6961 gatcaccatc atggaaagaa gcagcttcga gaagaatccc atcgactttc tggaagccaa
     7021 gggctacaaa gaagtgaaaa aggacctgat catcaagctg cctaagtact ccctgttcga
     7081 gctggaaaac ggccggaaga gaatgctggc ctctgccggc gaactgcaga agggaaacga
     7141 actggccctg ccctccaaat atgtgaactt cctgtacctg gccagccact atgagaagct
     7201 gaagggctcc cccgaggata atgagcagaa acagctgttt gtggaacagc acaagcacta
     7261 cctggacgag atcatcgagc agatcagcga gttctccaag agagtgatcc tggccgacgc
     7321 taatctggac aaagtgctgt ccgcctacaa caagcaccgg gataagccca tcagagagca
     7381 ggccgagaat atcatccacc tgtttaccct gaccaatctg ggagcccctg ccgccttcaa
     7441 gtactttgac accaccatcg accggaagag gtacaccagc accaaagagg tgctggacgc
     7501 caccctgatc caccagagca tcaccggcct gtacgagaca cggatcgacc tgtctcagct
     7561 gggaggcgac aaaaggccgg cggccacgaa aaaggccggc caggcaaaaa agaaaaaggg
     7621 cagtggagag ggcagaggaa gtctgctaac atgcggtgac gtcgaggaga atcctggccc
     7681 aatggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa cggctacaat
     7741 caacagcatc cccatctctg aagactacag cgtcgccagc gcagctctct ctagcgacgg
     7801 ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg cagaactcgt
     7861 ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg
     7921 aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc ttctcgatct
     7981 gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg cagttgggat
     8041 tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa aaacgttgtt gtttggggtt
     8101 gaattactct tctagaccgc ggcgcgcccg ataatcaacc tctggattac aaaatttgtg
     8161 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     8221 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     8281 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     8341 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     8401 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     8461 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     8521 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg
     8581 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     8641 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     8701 ccctttgggc cgcctccccg catcgatacc gtcgacctcg atcgagacct agaaaaacat
     8761 ggagcaatca caagtagcaa tacagcagct accaatgctg attgtgcctg gctagaagca
     8821 caagaggagg aggaggtggg ttttccagtc acacctcagg tacctttaag accaatgact
     8881 tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcta
     8941 attcactccc aacgaagaca agatatcctt gatctgtgga tctaccacac acaaggctac
     9001 ttccctgatt ggcagaacta cacaccaggg ccagggatca gatatccact gacctttgga
     9061 tggtgctaca agctagtacc agttgagcaa gagaaggtag aagaagccaa tgaaggagag
     9121 aacacccgct tgttacaccc tgtgagcctg catgggatgg atgacccgga gagagaagta
     9181 ttagagtgga ggtttgacag ccgcctagca tttcatcaca tggcccgaga gctgcatccg
     9241 gactgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
     9301 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
     9361 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
     9421 atctctagca gcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     9481 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     9541 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     9601 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     9661 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     9721 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     9781 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     9841 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     9901 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     9961 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
    10021 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
    10081 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
    10141 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
    10201 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
    10261 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
    10321 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
    10381 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
    10441 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
    10501 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
    10561 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
    10621 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
    10681 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
    10741 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
    10801 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
    10861 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
    10921 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
    10981 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
    11041 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
    11101 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
    11161 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
    11221 tttccccgaa aagtgccacc tgac
//
