LOCUS Exported 18664 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Target a Cre-dependent GCaMP6s cassette and a tdTomato-P2A-tTA2 cassette to the mouse TIGRE locus. ACCESSION . VERSION . KEYWORDS Ai163 (TIT2L-GC6s-ICL-TPT) targeting vector SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 18664) AUTHORS Daigle TL, Madisen L, Hage TA, Valley MT, Knoblich U, Larsen RS, Takeno MM, Huang L, Gu H, Larsen R, Mills M, Bosma-Moody A, Siverts LA, Walker M, Graybuck LT, Yao Z, Fong O, Nguyen TN, Garren E, Lenz GH, Chavarha M, Pendergraft J, Harrington J, Hirokawa KE, Harris JA, Nicovich PR, McGraw MJ, Ollerenshaw DR, Smith KA, Baker CA, Ting JT, Sunkin SM, Lecoq J, Lin MZ, Boyden ES, Murphy GJ, da Costa NM, Waters J, Li L, Tasic B, Zeng H TITLE A Suite of Transgenic Driver and Reporter Mouse Lines with Enhanced Brain-Cell-Type Targeting and Functionality. JOURNAL Cell. 2018 Jul 12;174(2):465-480.e22. doi: 10.1016/j.cell.2018.06.035. PUBMED 30007418 REFERENCE 2 (bases 1 to 18664) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..18664 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(29..51) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 151..170 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 364..386 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 378..395 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 379..395 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" polyA_signal 2975..3056 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind 3000..3019 /label=SV40pA-R /note="SV40 polyA, reverse primer" protein_bind 3070..3340 /label=tetracycline response element /bound_moiety="tetracycline repressor TetR" /note="contains seven copies of the tetracycline operator tetO" promoter 3373..3411 /label=minimal CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 3408..3432 /label=LNCX /note="Human CMV promoter, forward primer" protein_bind 3545..3578 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." polyA_signal 3655..3703 /note="synthetic polyadenylation signal" polyA_signal 3712..4188 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" polyA_signal 4200..4661 /label=PGK poly(A) signal /note="mouse phosphoglycerate kinase 1 polyadenylation signal" protein_bind 4688..4721 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." regulatory 4732..4741 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4738..6090 /codon_start=1 /product="improved fluorescent protein-based calcium sensor (Chen et al., 2013)" /label=GCaMP6s /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK" regulatory 4834..4843 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind 5124..5145 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(5251..5272) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(5512..5531) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" misc_feature 6115..6703 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" CDS complement(6586..6597) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 6734..6941 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" enhancer 9465..9768 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 9770..10047 /label=chicken beta-actin promoter intron 10048..11064 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 10988..11010 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 11072..11091 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind complement(11152..11185) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." polyA_signal 11223..11271 /note="synthetic polyadenylation signal" polyA_signal 11280..11756 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(11791..11810) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 11835..11883 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" protein_bind complement(12057..12090) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." regulatory 12107..12116 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind complement(12246..12264) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 12514..12533 /label=mCherry-F /note="mCherry, forward primer" primer_bind 12708..12731 /label=DsRed1-C /note="DsRed1, forward primer" CDS 13550..13606 /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 13607..14353 /codon_start=1 /product="improved tetracycline-controlled transactivator" /label=tTA-Advanced /note="In the Tet-Off(R) system, tTA-Advanced binds to the Tet-responsive element and stimulates transcription only in the absence of tetracycline or doxycycline." /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA DALDDFDLDMLPADALDDFDLDMLPG" misc_feature 14396..14984 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" CDS complement(14867..14878) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 15015..15222 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" protein_bind 15236..15269 /label=attB /bound_moiety="phage phi-C31 integrase" /note="minimal attB site for the phi-C31 integrase (Groth et al., 2000)" promoter 15296..15795 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind complement(15331..15354) /label=MSCV-rev /note="Murine stem cell virus, reverse primer" primer_bind 15728..15747 /label=mPGK-F /note="Mouse PGK promoter, forward primer" primer_bind complement(16443..16459) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(16443..16459) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(16456..16478) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 16467..16483 /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(16491..16521) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 16536..16557 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(16674..16691) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(16845..17433) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(16925..16944) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(17604..18464) /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 18227..18246 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(18465..18569) /gene="bla" /label=AmpR promoter primer_bind 18637..18655 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca 61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg 121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc 181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt accgaagttc 421 ctattcttca aatagtatag gaacttcttt cggccggcca atcgattgga tccgtttaat 481 tcgagctcgc tagagcccca tcctcactga ctccgtcctg gagttggatg agagataatg 541 gccttacgtt gtgccagggg agggtcgggc tggatttagc aagatttacc ttctccaaag 601 agcggtgctg cagtggcaca gctgcccacg gaggtggggg ggtcaccgtc cctggaggtg 661 atgaagaact gtggggatgt ggcactgagg gacatggcca gtgggcacgg tgggtgggtt 721 ggggttggtc ttggggatct tggagggctt ttccagcctt catgatttga cgattgtatg 781 aacatctaca tggcaattct ccagctgcct gtcccagtcc tactgaccca gctgtatctc 841 tccaggcaag ctcttccacc ccttctgctt gcatccagac accatcaaac atgcaggctc 901 agacacaggg accagcagtg tctgtggcct ttttgtgctc ctctccatgc tgggttttaa 961 cttgctcttt gtccttctat cctatcttct tatccttaag gctgttctga acgctgtgac 1021 ttggagagtg tcccagagcc ctcaacacct gcatgtccca cgtccatgct gtcctgcact 1081 tccttatccc caagatctgc ctctccgtga tgcactgaat tggcaaacat gtgtcacccc 1141 agaccaacaa tgtcacagca aactccccct tgataggaca agggggaatg gctttacact 1201 gagacagggg aggtttgggt tggatatgag gaggcagttt ttcccccaga gggtggtgac 1261 gcactgaaca ggttgcccaa ggaggctgtg gatgccccat ccctgcaggc attcaaggcc 1321 aggctggatg tggctctggg cagcctgggc tgctggttga tgaccctgca catagcaggg 1381 ggttggatct ggatgagcac tgtgctcctt tgcaacccag gccgttctat gattctgtca 1441 ttctaaatct ctctttcagc ctaaagcttt ttccccgtat ccccccaggt gtctgcaggc 1501 tcaaagagca gcgagaagcg ttcagaggaa agcgatcccg tgccaccttc cccgtgcccg 1561 ggctgtcccc gcacgctgcc ggctcgggga tgcgggggga gcgccggacc ggagcggagc 1621 cccgggcggc tcgctgctgc cccctagcgg gggagggacg taattacatc cctgggggct 1681 ttgggggggg gctgtccctc tagcgagctc ggtacccaga gccccatcct cactgactcc 1741 gtcctggagt tggatgagag ataatggcct tacgttgtgc caggggaggg tcgggctgga 1801 tttagcaaga tttaccttct ccaaagagcg gtgctgcagt ggcacagctg cccacggagg 1861 tgggggggtc accgtccctg gaggtgatga agaactgtgg ggatgtggca ctgagggaca 1921 tggccagtgg gcacggtggg tgggttgggg ttggtcttgg ggatcttgga gggcttttcc 1981 agccttcatg atttgacgat tgtatgaaca tctacatggc aattctccag ctgcctgtcc 2041 cagtcctact gacccagctg tatctctcca ggcaagctct tccacccctt ctgcttgcat 2101 ccagacacca tcaaacatgc aggctcagac acagggacca gcagtgtctg tggccttttt 2161 gtgctcctct ccatgctggg ttttaacttg ctctttgtcc ttctatccta tcttcttatc 2221 cttaaggctg ttctgaacgc tgtgacttgg agagtgtccc agagccctca acacctgcat 2281 gtcccacgtc catgctgtcc tgcacttcct tatccccaag atctgcctct ccgtgatgca 2341 ctgaattggc aaacatgtgt caccccagac caacaatgtc acagcaaact cccccttgat 2401 aggacaaggg ggaatggctt tacactgaga caggggaggt ttgggttgga tatgaggagg 2461 cagtttttcc cccagagggt ggtgacgcac tgaacaggtt gcccaaggag gctgtggatg 2521 ccccatccct gcaggcattc aaggccaggc tggatgtggc tctgggcagc ctgggctgct 2581 ggttgatgac cctgcacata gcagggggtt ggatctggat gagcactgtg ctcctttgca 2641 acccaggccg ttctatgatt ctgtcattct aaatctctct ttcagcctaa agctttttcc 2701 ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga gaagcgttca gaggaaagcg 2761 atcccgtgcc accttccccg tgcccgggct gtccccgcac gctgccggct cggggatgcg 2821 gggggagcgc cggaccggag cggagccccg ggcggctcgc tgctgccccc tagcggggga 2881 gggacgtaat tacatccctg ggggctttgg gggggggctg tccctctagg ggtacccggg 2941 gatccaccgg tatcgattcg gccggccgac gtcggcagtg aaaaaaatgc tttatttgtg 3001 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagtttcga 3061 gtttaccact ccctatcagt gatagagaaa agtgaaagtc gagtttacca ctccctatca 3121 gtgatagaga aaagtgaaag tcgagtttac cactccctat cagtgataga gaaaagtgaa 3181 agtcgagttt accactccct atcagtgata gagaaaagtg aaagtcgagt ttaccactcc 3241 ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt gatagagaaa 3301 agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagctcgg 3361 tacccgggtc gaggtaggcg tgtacggtgg gaggcctata taagcagagc tcgtttagtg 3421 aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg 3481 gaccgatcca gcctccgcgg ccccgaattc gagctcggta cccggggatc aaattcaatt 3541 aaatataact tcgtatagca tacattatac gaagttattc gcgatgaata aatgaaagct 3601 tgcagatccc ctcagcacca tggctagcgg cagcctcgga gtttgaatag atagaataaa 3661 atatctttat tttcattcca tctgtgtgtt ggttttttgt gtgagatcta cgggtggcat 3721 ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc agtgcccacc 3781 agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc cttctataat 3841 attatggggt ggaggggggt ggtatggagc aaggggcaag ttgggaagac aacctgtagg 3901 gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg gctcactgca 3961 atctccgcct cctgggttca agcgattctc ctgcctcagg ctcccgagtt gttgggattc 4021 caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg gggtttcacc 4081 atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc ttggcctccc 4141 aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct gcctcagcgc 4201 tgtaagtctg cagaaattga tgatctatta aacaataaag atgtccacta aaatggaagt 4261 ttttcctgtc atactttgtt aagaagggtg agaacagagt acctacattt tgaatggaag 4321 gattggagct acgggggtgg gggtggggtg ggattagata aatgcctgct ctttactgaa 4381 ggctctttac tattgcttta tgataatgtt tcatagttgg atatcataat ttaaacaagc 4441 aaaaccaaat taagggccag ctcattcctc ccactcatga tctatagatc tatagatctc 4501 tcgtgggatc attgtttttc tcttgattcc cactttgtgg ttctaagtac tgtggtttcc 4561 aaatgtgtca gtttcatagc ctgaagaacg agatcagcag cctctgttcc acatacactt 4621 cattctcagt attgttttgc caagttctaa ttccatcaga aagcttttaa ttaactgaat 4681 agctaacata acttcgtata gcatacatta tacgaagtta tcacgcgtgc cgccaccatg 4741 ggttctcatc atcatcatca tcatggtatg gctagcatga ctggtggaca gcaaatgggt 4801 cgggatctgt acgacgatga cgataaggat ctcgccacca tggtcgactc atcacgtcgt 4861 aagtggaata agacaggtca cgcagtcaga gctataggtc ggctgagctc actcgagaac 4921 gtctatatca aggccgacaa gcagaagaac ggcatcaagg cgaacttcca catccgccac 4981 aacatcgagg acggcggcgt gcagctcgcc taccactacc agcagaacac ccccatcggc 5041 gacggccccg tgctgctgcc cgacaaccac tacctgagcg tgcagtccaa actttcgaaa 5101 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc 5161 actctcggca tggacgagct gtacaagggc ggtaccggag ggagcatggt gagcaagggc 5221 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc 5281 cacaagttca gcgtgtccgg cgagggtgag ggcgatgcca cctacggcaa gctgaccctg 5341 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg 5401 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc 5461 aagtccgcca tgcccgaagg ctacatccag gagcgcacca tcttcttcaa ggacgacggc 5521 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag 5581 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac 5641 ctgccggacc aactgactga agagcagatc gcagaattta aagaggcttt ctccctattt 5701 gacaaggacg gggatgggac aataacaacc aaggagctgg ggacggtgat gcggtctctg 5761 gggcagaacc ccacagaagc agagctgcag gacatgatca atgaagtaga tgccgacggt 5821 gacggcacaa tcgacttccc tgagttcctg acaatgatgg caagaaaaat gaaatacagg 5881 gacacggaag aagaaattag agaagcgttc ggtgtgtttg ataaggatgg caatggctac 5941 atcagtgcag cagagcttcg ccacgtgatg acaaaccttg gagagaagtt aacagatgaa 6001 gaggttgatg aaatgatcag ggaagcagac atcgatgggg atggtcaggt aaactacgaa 6061 gagtttgtac aaatgatgac agcgaagtaa acgcgttacg taaagcttgt cgataatcaa 6121 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt 6181 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct 6241 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc 6301 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg 6361 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc 6421 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc 6481 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctat 6541 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca 6601 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt 6661 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgaca ccgagcgctg 6721 ctcgaggcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg 6781 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt 6841 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc 6901 aagggggagg attgggaaga caatagcagg catgctgggg acacgtggga tcctctagag 6961 ccccatcctc actgactccg tcctggagtt ggatgagaga taatggcctt acgttgtgcc 7021 aggggagggt cgggctggat ttagcaagat ttaccttctc caaagagcgg tgctgcagtg 7081 gcacagctgc ccacggaggt gggggggtca ccgtccctgg aggtgatgaa gaactgtggg 7141 gatgtggcac tgagggacat ggccagtggg cacggtgggt gggttggggt tggtcttggg 7201 gatcttggag ggcttttcca gccttcatga tttgacgatt gtatgaacat ctacatggca 7261 attctccagc tgcctgtccc agtcctactg acccagctgt atctctccag gcaagctctt 7321 ccaccccttc tgcttgcatc cagacaccat caaacatgca ggctcagaca cagggaccag 7381 cagtgtctgt ggcctttttg tgctcctctc catgctgggt tttaacttgc tctttgtcct 7441 tctatcctat cttcttatcc ttaaggctgt tctgaacgct gtgacttgga gagtgtccca 7501 gagccctcaa cacctgcatg tcccacgtcc atgctgtcct gcacttcctt atccccaaga 7561 tctgcctctc cgtgatgcac tgaattggca aacatgtgtc accccagacc aacaatgtca 7621 cagcaaactc ccccttgata ggacaagggg gaatggcttt acactgagac aggggaggtt 7681 tgggttggat atgaggaggc agtttttccc ccagagggtg gtgacgcact gaacaggttg 7741 cccaaggagg ctgtggatgc cccatccctg caggcattca aggccaggct ggatgtggct 7801 ctgggcagcc tgggctgctg gttgatgacc ctgcacatag cagggggttg gatctggatg 7861 agcactgtgc tcctttgcaa cccaggccgt tctatgattc tgtcattcta aatctctctt 7921 tcagcctaaa gctttttccc cgtatccccc caggtgtctg caggctcaaa gagcagcgag 7981 aagcgttcag aggaaagcga tcccgtgcca ccttccccgt gcccgggctg tccccgcacg 8041 ctgccggctc ggggatgcgg ggggagcgcc ggaccggagc ggagccccgg gcggctcgct 8101 gctgccccct agcgggggag ggacgtaatt acatccctgg gggctttggg ggggggctgt 8161 ccctctagag ccccatcctc actgactccg tcctggagtt ggatgagaga taatggcctt 8221 acgttgtgcc aggggagggt cgggctggat ttagcaagat ttaccttctc caaagagcgg 8281 tgctgcagtg gcacagctgc ccacggaggt gggggggtca ccgtccctgg aggtgatgaa 8341 gaactgtggg gatgtggcac tgagggacat ggccagtggg cacggtgggt gggttggggt 8401 tggtcttggg gatcttggag ggcttttcca gccttcatga tttgacgatt gtatgaacat 8461 ctacatggca attctccagc tgcctgtccc agtcctactg acccagctgt atctctccag 8521 gcaagctctt ccaccccttc tgcttgcatc cagacaccat caaacatgca ggctcagaca 8581 cagggaccag cagtgtctgt ggcctttttg tgctcctctc catgctgggt tttaacttgc 8641 tctttgtcct tctatcctat cttcttatcc ttaaggctgt tctgaacgct gtgacttgga 8701 gagtgtccca gagccctcaa cacctgcatg tcccacgtcc atgctgtcct gcacttcctt 8761 atccccaaga tctgcctctc cgtgatgcac tgaattggca aacatgtgtc accccagacc 8821 aacaatgtca cagcaaactc ccccttgata ggacaagggg gaatggcttt acactgagac 8881 aggggaggtt tgggttggat atgaggaggc agtttttccc ccagagggtg gtgacgcact 8941 gaacaggttg cccaaggagg ctgtggatgc cccatccctg caggcattca aggccaggct 9001 ggatgtggct ctgggcagcc tgggctgctg gttgatgacc ctgcacatag cagggggttg 9061 gatctggatg agcactgtgc tcctttgcaa cccaggccgt tctatgattc tgtcattcta 9121 aatctctctt tcagcctaaa gctttttccc cgtatccccc caggtgtctg caggctcaaa 9181 gagcagcgag aagcgttcag aggaaagcga tcccgtgcca ccttccccgt gcccgggctg 9241 tccccgcacg ctgccggctc ggggatgcgg ggggagcgcc ggaccggagc ggagccccgg 9301 gcggctcgct gctgccccct agcgggggag ggacgtaatt acatccctgg gggctttggg 9361 ggggggctgt ccctctagag tcgacggcgc gccctgcagg aatctagtta ttaatagtaa 9421 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg 9481 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg 9541 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta 9601 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt 9661 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac 9721 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc 9781 cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta 9841 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gcgcgcgccr 9901 ggsggggsgg ggsggggsgr ggggsggggc ggggcgaggc ggagaggtgc ggcggcagcc 9961 aatcagagcg gcgcgctccg aaagtttcct tttatggcga ggcggcggcg gcggcggccc 10021 tataaaaagc gaagcgcgcg gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc 10081 cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt actcccacag 10141 gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agcgcttggt ttaatgacgg 10201 cttgtttctt ttctgtggct gcgtgaaagc cttgaggggc tccgggaggg ccctttgtgc 10261 ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc cgcgtgcggc 10321 tccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg tgcgctccgc 10381 agtgtgcgcg aggggagcgc ggccgggggc ggtgccccgc ggtgcggggg gggctgcgag 10441 gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg tgtgggcgcg 10501 tcggtcgggc tgcaaccccc cctgcacccc cctccccgag ttgctgagca cggcccggct 10561 tcgggtgcgg ggctccgtac ggggcgtggc gcggggctcg ccgtgccggg cggggggtgg 10621 cggcaggtgg gggtgccggg cggggcgggg ccgcctcggg ccggggaggg ctcgggggag 10681 gggcgcggcg gcccccggag cgccggcggc tgtcgaggcg cggcgagccg cagccattgc 10741 cttttatggt aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat ctgtgcggag 10801 ccgaaatctg ggaggcgccg ccgcaccccc tctagcgggc gcggggcgaa gcggtgcggc 10861 gccggcagga aggaaatggg cggggagggc cttcgtgcgt cgccgcgccg ccgtcccctt 10921 ctccctctcc agcctcgggg ctgtccgcgg ggggacggct gccttcgggg gggacggggc 10981 agggcggggt tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc taaccatgtt 11041 catgccttct tctttttcct acagctcctg ggcaacgtgc tggttattgt gctgtctcat 11101 cattttggca aagaattgat ttgataccgc gggccccgcg tggacggtac cataacttcg 11161 tataaagtat cctatacgaa gttatccatg gctagcggca gcctcggagt ttgaatagat 11221 agaataaaat atctttattt tcattccatc tgtgtgttgg ttttttgtgt gagatctacg 11281 ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag 11341 tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct 11401 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa 11461 cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc 11521 tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcaggct cccgagttgt 11581 tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg 11641 gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt 11701 ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttctgc 11761 ctcagctaat tgagtagggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg 11821 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacgggtgtt gggtcgtttg 11881 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc 11941 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg 12001 aaggcccagg gctcgcagcc aacgtcgggg cggcaggcct cgcgagggtt tctagaataa 12061 cttcgtataa agtatcctat acgaagttat acgcgcgcgt actagcgcca ccatggtgag 12121 caagggcgag gaggtcatca aagagttcat gcgcttcaag gtgcgcatgg agggctccat 12181 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca 12241 gaccgccaag ctgaaggtga ccaagggcgg ccccctgccc ttcgcctggg acatcctgtc 12301 cccccagttc atgtacggct ccaaggcgta cgtgaagcac cccgccgaca tccccgatta 12361 caagaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg 12421 cggtctggtg accgtgaccc aggactcctc cctgcaggac ggcacgctga tctacaaggt 12481 gaagatgcgc ggcaccaact tcccccccga cggccccgta atgcagaaga agaccatggg 12541 ctgggaggcc tccaccgagc gcctgtaccc ccgcgacggc gtgctgaagg gcgagatcca 12601 ccaggccctg aagctgaagg acggcggcca ctacctggtg gagttcaaga ccatctacat 12661 ggccaagaag cccgtgcaac tgcccggcta ctactacgtg gacaccaagc tggacatcac 12721 ctcccacaac gaggactaca ccatcgtgga acagtacgag cgctccgagg gccgccacca 12781 cctgttcctg gggcatggta ccggcagcac cggcagcggc agctccggca ccgcctcctc 12841 cgaggacaac aacatggccg tgattaagga gtttatgagg tttaaagtca ggatggaggg 12901 cagcatgaac gggcacgagt ttgagattga aggagagggg gaagggaggc cctatgaggg 12961 cacccagaca gcaaagctga aagtgacaaa gggcggccct ctgccattcg catgggacat 13021 cctgtctcct cagtttatgt atggaagtaa agcctacgtg aaacaccctg ctgacattcc 13081 agattacaag aaactgagct tcccagaggg gtttaagtgg gaaagagtga tgaacttcga 13141 agacgggggc ctggtgactg tcacccagga cagctctctg caggacggca ctctgatcta 13201 taaggtgaaa atgcgcggca ccaattttcc ccctgatggc ccagtcatgc agaagaagac 13261 aatgggctgg gaggcttcaa cagaaaggct gtaccccaga gacggcgtgc tgaagggaga 13321 gatccaccag gcactgaagc tgaaagatgg agggcattat ctggtcgaat tcaaaaccat 13381 ctacatggcc aagaaacccg tgcagctgcc tggctactat tacgtcgaca caaagctgga 13441 tattactagc cataacgaag actacaccat tgtggagcag tatgagcgga gcgaggggag 13501 gcaccatctg tttctgtacg gcatggacga gctgtacaag ggaagcggag ctactaactt 13561 cagcctgctg aagcaggctg gagacgtgga ggagaaccct ggacctatgt ctagactgga 13621 caagagcaaa gtcataaact ctgctctgga attactcaat gaagtcggta tcgaaggcct 13681 gacgacaagg aaactcgctc aaaagctggg agttgagcag cctaccctgt actggcacgt 13741 gaagaacaag cgggccctgc tcgatgccct ggcaatcgag atgctggaca ggcatcatac 13801 ccacttctgc cccctggaag gcgagtcatg gcaagacttt ctgcggaaca acgccaagtc 13861 attccgctgt gctctcctct cacatcgcga cggggctaaa gtgcatctcg gcacccgccc 13921 aacagagaaa cagtacgaaa ccctggaaaa tcagctcgcg ttcctgtgtc agcaaggctt 13981 ctccctggag aacgcactgt acgctctgtc cgccgtgggc cactttacac tgggctgcgt 14041 attggaggat caggagcatc aagtagcaaa agaggaaaga gagacaccta ccaccgattc 14101 tatgccccca cttctgagac aagcaattga gctgttcgac catcagggag ccgaacctgc 14161 cttccttttc ggcctggaac taatcatatg tggcctggag aaacagctaa agtgcgaaag 14221 cggcgggccg gccgacgccc ttgacgattt tgacttagac atgctcccag ccgatgccct 14281 tgacgacttt gaccttgata tgctgcctgc tgacgctctt gacgattttg accttgacat 14341 gctccccggg taactaagta aggatcaaat ggcgcgttac gtaaagcttg tcgataatca 14401 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt 14461 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc 14521 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc 14581 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg 14641 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc 14701 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg 14761 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta 14821 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc 14881 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct 14941 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgac accgagcgct 15001 gctcgaggcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt 15061 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat 15121 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag 15181 caagggggag gattgggaag acaatagcag gcatgctggg gacacgtggg atccggtgcc 15241 agggcgtgcc cttgggctcc ccgggcgcgt ttgggcggcc gcatcgaatt ctaccgggta 15301 ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcagcccc gctgggcact 15361 tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa 15421 ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag 15481 ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc 15541 actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc 15601 agcggccaat agcagctttg ctccttcgct ttctgggctc agaggctggg aaggggtggg 15661 tccgggggcg ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga 15721 ggcccggcat tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca 15781 tctccgggcc tttcgacctg caatcgccgc tagccgccac catgaaaaag cctgaactca 15841 ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc 15901 agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg 15961 tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact 16021 ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc 16081 tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg 16141 aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc 16201 ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat 16261 ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg 16321 acgacaccgt cagtgtaagt gctagcccct aggcctgaat cgaagttcct attcttcaaa 16381 aggtatagga acttcgagct cgtcgactgc agaggcctgc atgcaagctt ggcgtaatca 16441 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga 16501 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt 16561 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga 16621 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc 16681 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg 16741 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc 16801 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc 16861 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga 16921 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc 16981 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat 17041 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg 17101 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc 17161 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga 17221 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact 17281 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt 17341 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag 17401 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg 17461 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa 17521 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata 17581 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg 17641 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata 17701 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg 17761 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct 17821 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt 17881 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc 17941 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga 18001 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt 18061 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc 18121 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa 18181 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca 18241 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca 18301 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct 18361 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc 18421 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa 18481 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt 18541 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc 18601 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt 18661 cgtc //