LOCUS Exported 13048 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Donor construct for insertion of NGN2 and BRN3A into the CLYBL safe harbor site. This is an all-in-one doxycycline-inducible construct, enabling iPSC differentiation into peripheral sensory neurons.. ACCESSION . VERSION . KEYWORDS pUCM-CLYBL-NGN2-BRN3A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13048) AUTHORS Nickolls AR, Lee MM, Espinoza DF, Szczot M, Lam RM, Wang Q, Beers J, Zou J, Nguyen MQ, Solinski HJ, AlJanahi AA, Johnson KR, Ward ME, Chesler AT, Bonnemann CG TITLE Transcriptional Programming of Human Mechanosensory Neuron Subtypes from Pluripotent Stem Cells. JOURNAL Cell Rep. 2020 Jan 21;30(3):932-946.e7. doi: 10.1016/j.celrep.2019.12.062. PUBMED 31968264 REFERENCE 2 (bases 1 to 13048) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..13048 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 266..285 /label=pBR322ori-F /note="pBR322 origin, forward primer" protein_bind 1391..1409 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1427..1445 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1463..1481 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1499..1517 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1535..1553 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1571..1589 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1607..1625 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind 1670..1694 /label=LNCX /note="Human CMV promoter, forward primer" CDS 2620..2673 /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 2977..3003 /codon_start=1 /product="9xHis affinity tag" /label=9xHis /translation="HHHHHHHHH" polyA_signal 3962..4083 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3999..4018) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4053..4072 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4132..4148) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4132..4148) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4145..4167) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4156..4172 /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(4180..4210) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4225..4246 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4363..4380) /label=L4440 /note="L4440 vector, forward primer" primer_bind 4672..4691 /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" polyA_signal 4690..4745 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind 4791..4810 /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" CDS complement(4901..5647) /codon_start=1 /product="modified rtTA protein that binds tightly to promoters containing the tet operator in the presence of doxycycline" /label=Tet-On(R) 3G /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA DALDDFDLDMLPADALDDFDLDMLPG" primer_bind complement(5679..5698) /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" intron 5706..6723 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind complement(5760..5782) /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" promoter complement(6725..7002) /label=chicken beta-actin promoter enhancer 7004..7383 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" polyA_signal 7389..7613 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind 7602..7619 /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" protein_bind complement(7635..7668) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind 8322..8341 /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" CDS complement(8342..8395) /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 complement(8417..9133) /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS complement(8417..9133) /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(8459..8480) /label=EGFP-C /note="EGFP, forward primer" primer_bind 8808..8827 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 9067..9088 /label=EGFP-N /note="EGFP, reverse primer" regulatory 9130..9139 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" LTR 9167..9435 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from human T-cell leukemia virus (HTLV) type 1" promoter complement(9448..9659) /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" misc_feature 9692..9783 /label=pause site /note="RNA polymerase II transcriptional pause signal from the human alpha-2 globin gene" primer_bind complement(9724..9743) /label=RVprimer3 /note="pGL3 vector, forward primer" polyA_signal 9797..9845 /note="synthetic polyadenylation signal" CDS complement(10605..10718) /codon_start=1 /product="streptavidin-binding peptide" /label=SBP /note="selected from a peptide library; binds streptavidin with nanomolar affinity (Keefe et al., 2001)" /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP" CDS complement(10800..10853) /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" misc_feature 10877..10902 /label=SA /note="splice acceptor site" protein_bind complement(10903..10936) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS 11794..12654 /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(12012..12031) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 12825..365 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" ORIGIN 1 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac 61 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 121 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg 181 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag 241 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt 301 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat 361 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc gaattcgccc ttagagcaga 421 gccaggaacc cctgtaggga aggggcagga gagccagggg catgagatgg tggacgagga 481 agggggacag ggaagcctga gcgcctctcc tgggcttgcc aaggactcaa acccagaagc 541 ccagagcagg gccttaggga agcgggacgg ctctgttgga gagtccagta ttgaattagt 601 gatagataca cagaaaaaaa gcaaattgaa taggtgaatg gataaacaaa ctgtggaaca 661 tccagacaat gaaatattca ccaataaaaa gaaatgagct atcaagccac attggctatg 721 cccttcacat ttaaggagga accttaaatg catattgcca aattttaaaa gctagtctga 781 aaatgctaca gactgtatga ttccaattat atgacatttt agaaaaggca aaactataga 841 gatcagtggt tgccagcggc tcatggaggc aggatgaatt ggtggaacac agggatgctt 901 tcagggtaat gaaaatatct tgtgtgatcc tgtaataatg gatacgtggc ataatgcatt 961 tgttaaaacc tgtaaactag ccaggtatag tggctttttt gcacctatag tctcagctac 1021 ttggtaggct gagacaggaa gatcacttga gcccaggact tcaagaccag cctgggtaat 1081 atagtaagac cccatctcta ttttttaaag aaattaagaa agaaaagaaa cccatcaact 1141 gtataacaca aagaatcgta atgtaaacta cagtcctgag taatgaaaca tttttaaaga 1201 aataaataaa ttcaaggcaa ttattaactt caggaaaaac agtaaagtaa acataagtat 1261 acaaatcaaa catggctcag ttgtgaaaat atttacaagg tcactttaag taacattgaa 1321 tgttgatata atcccaaaat atatttatgt tggaaggatg aggagatatc tccaccccac 1381 agtggtttac tccctatcag tgatagagaa cgtatgaaga gtttactccc tatcagtgat 1441 agagaacgta tgcagacttt actccctatc agtgatagag aacgtataag gagtttactc 1501 cctatcagtg atagagaacg tatgaccagt ttactcccta tcagtgatag agaacgtatc 1561 tacagtttac tccctatcag tgatagagaa cgtatatcca gtttactccc tatcagtgat 1621 agagaacgta taagctttag gcgtgtacgg tgggcgccta taaaagcaga gctcgtttag 1681 tgaaccgtca gatcgcctgg agcaattcca caacactttt gtcttatacc aactttccgt 1741 accacttcct accctcgtaa acttaaggtt aattaagcca ccatgttcgt gaagtccgag 1801 acactggaac tgaaagagga agaggacgtg ctggtgctgc tgggctctgc ctctcctgct 1861 ctggctgctc tgacacccct gagcagcagc gccgacgagg aagaagaaga ggaacctggc 1921 gctagcggcg gagccagaag gcagagaggc gctgaagctg gacagggcgc aagaggcgga 1981 gtggctgctg gcgcagaagg atgcagacct gccagactgc tgggcctggt gcacgactgc 2041 aagagaaggc cctctagagc cagagccgtg tccagaggcg ccaagacagc cgagacagtg 2101 cagcggatca agaaaaccag acggctgaag gccaacaacc gcgagcggaa ccggatgcac 2161 aacctgaacg ccgccctgga cgccctgaga gaggtgctgc ctaccttccc cgaggacgcc 2221 aagctgacca agatcgaaac cctgagattc gcccacaact acatctgggc cctgaccgag 2281 actctgcggc tggccgatca ttgtggcggc ggaggcggag gactgccagg cgctctgttt 2341 tctgaagccg tgctgctgtc tcctggcgga gcttctgctg ccctgtctag cagcggcgat 2401 tctcctagcc ctgccagcac ctggtcctgc accaatagcc cagcccctag ctccagcgtg 2461 tccagcaatt ccaccagccc ttacagctgc accctgagcc ctgcttctcc agccggcagc 2521 gacatggact actggcagcc tcccccccct gacaaacatc gctacgcccc ccatctgcct 2581 atcgcccggg attgcatcat gcacagcaga ggctctggcg agggcagagg cagcctgctg 2641 acatgtggcg acgtggaaga gaaccctggc cccggatcca tgatgtccat gaactccaaa 2701 cagccacact ttgccatgca cccaactctg cctgagcata agtacccatc cctgcattcc 2761 tcctccgagg caatcaggag ggcttgtctg cctactcctc ctctccagag caacctgttc 2821 gcttctctgg acgaaacact gctcgcaaga gccgaagccc tcgcagccgt ggacattgct 2881 gtttctcagg gcaagtccca tcctttcaaa cccgacgcca cctaccacac catgaatagc 2941 gtcccatgca ccagcacctc cactgtccca ctcgctcatc atcatcacca tcaccaccac 3001 caccaggcac tggagcctgg tgatctcctc gaccacatct cctctccatc cctggctctg 3061 atggctggag ctggaggtgc tggagcagct gcaggtggag gtggagccca cgacggtcca 3121 ggaggaggtg gaggaccagg aggaggagga ggacctggag gtggtcctgg tggtggagga 3181 ggaggaggac caggaggagg tggaggagga ccaggaggag gactcctcgg aggaagcgct 3241 caccctcatc ctcacatgca cagcctgggt cacctgtccc accctgctgc tgctgctgct 3301 atgaacatgc cttctggact gcctcaccct ggcctggttg cagcagctgc tcatcacggt 3361 gcagctgctg cagctgcagc tgctgctgca ggtcaggtcg ctgctgcctc tgctgcagct 3421 gccgtcgttg gagctgctgg actggcctcc atctgcgatt ccgacacaga cccaagagaa 3481 ctcgaagcat tcgctgaaag attcaaacaa agaagaatca aactcggtgt tactcaagca 3541 gatgtgggaa gcgcactggc taacctcaag atccctggag tcggcagcct gagccagtcc 3601 accatctgtc gctttgaatc tctgaccctg tcccacaaca acatgattgc cctcaagcct 3661 atcctccagg cttggctgga ggaggcagag ggagcacaac gcgagaagat gaacaagcca 3721 gagctgttca acggaggcga gaagaagcgc aagcgcactt ccattgctgc acctgagaag 3781 agatccctgg aggcatactt cgcagtgcag cccaggccat cttctgagaa gatcgcagct 3841 attgccgaga agctcgacct caagaagaac gtggtcaggg tgtggttctg caatcagagg 3901 cagaaacaga agagaatgaa gttctccgca acctactaat gtacaatcga tcgcccaccc 3961 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac 4021 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc 4081 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct 4141 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat 4201 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc 4261 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg 4321 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct 4381 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggacg gccgcacatg 4441 taaggaaaat tttagggatg ttaaagaaaa aaataacaca aaacaaaata taaaaaaaat 4501 ctaacctcaa gtcaaggctt ttctatggaa taaggaatgg acagcagggg gctgtttcat 4561 atactgatga cctctttata gccaaccttt gttcatggca gccagcatat gggcatatgt 4621 tgccaaactc taaaccaaat actcattctg atgttttaaa tgatttgccc tcccatatgt 4681 ccttccgagt gagagacaca aaaaattcca acacactatt gcaatgaaaa taaatttcct 4741 ttattagcca gaagtcagat gctcaagggg cttcatgatg tccccataat ttttggcaga 4801 gggaaaaaga tctcagtggt atttgtgagc cagggcattg gccacaccag tcaccacctt 4861 ctgataggca gcctgcacct gaggagtgaa ttcacgcgtt ttagcctggc agcatatcga 4921 gatcgaagtc atcgagagcg tcagcaggga gcatatccag atcaaaatca tccagggcat 4981 cggcgggcag catgtccagg tcgaagtcgt ccagggcgtc ggtgggtccg ccgctctcgc 5041 acttcagctg cttctcgagg ccgcagatga tcagttccag gccgaacagg aaggcgggct 5101 cggcgccctg tctgtcgaac agctcgatgg cctgcttcag cagggggggc atgctgtcgg 5161 tggtgggtgt ctctctttcc tctttggcga cctggtgttc ctgttcttcc agcacgcagc 5221 ccagggtgaa gtggcccacg gcgctcaggg cgtacagggc gttttccagg ctgaagccct 5281 gctggcacag gaaggccagc tggttttcca gtgtctcgta ctgcttctcg gtgggtctgg 5341 tgcccaggtg cactttggcg ccgtcccggt ggctcagcag ggcgcatctg tagctcttgg 5401 cgttgttccg caggaagtcc tgccagctct cgccttccag agggcagctg tgggtgtggt 5461 gccggtccag catctcgatg ggcagggcgt ccagcagggc ccgcttgttc ttcacgtgcc 5521 agtacagggt gggctgttcc acgcccagtt tctgggccag cttccgggtg gtcaggccct 5581 cgatgcccac gccgttcagc agttccaggg cgctgttgat cactttgctc ttgtccagcc 5641 ggctcatgtc gacattcttt gccaaaatga tgagacagca caataaccag cacgttgccc 5701 aggagctgta ggaaaaagaa gaaggcatga acatggttag cagaggctct agagccgccg 5761 gtcacacgcc agaagccgaa ccccgccctg ccccgtcccc cccgaaggca gccgtccccc 5821 cgcggacagc cccgaggctg gagagggaga aggggacggc ggcgcggcga cgcacgaagg 5881 ccctccccgc ccatttcctt cctgccggcg ccgcaccgct tcgccccgcg cccgctagag 5941 ggggtgcggc ggcgcctccc agatttcggc tccgcacaga tttgggacaa aggaagtccc 6001 tgcgccctct cgcacgatta ccataaaagg caatggctgc ggctcgccgc gcctcgacag 6061 ccgccggcgc tccgggggcc gccgcgcccc tcccccgagc cctccccggc ccgaggcggc 6121 cccgccccgc ccggcacccc cacctgccgc caccccccgc ccggcacggc gagccccgcg 6181 ccacgccccg tacggagccc cgcacccgaa gccgggccgt gctcagcaac tcggggaggg 6241 gggtgcaggg ggggttgcag cccgaccgac gcgcccacac cccctgctca cccccccacg 6301 cacacacccc gcacgcagcc tttgttcccc tcgcagcccc ccccgcaccg cggggcaccg 6361 cccccggccg cgctcccctc gcgcacactg cggagcgcac aaagccccgc gccgcgcccg 6421 cagcgctcac agccgccggg cagcgcggag ccgcacgcgg cgctccccac gcacacacac 6481 acgcacgcac cccccgagcc gctccccccg cacaaagggc cctcccggag cccctcaagg 6541 ctttcacgca gccacagaaa agaaacaagc cgtcattaaa ccaagcgcta attacagccc 6601 ggaggagaag ggccgtcccg cccgctcacc tgtgggagta acgcggtcag tcagagccgg 6661 ggcgggcggc gcgaggcggc ggcggagcgg ggcacggggc gaaggcagcg tcgcagcgac 6721 tccccgcccg ccgcgcgctt cgctttttat agggccgccg ccgccgccgc ctcgccataa 6781 aaggaaactt tcggagcgcg ccgctctgat tggctgccgc cgcacctctc cgcctcgccc 6841 cgccccgccc ctcgccccgc cccgccccgc ctggcgcgcg cccccccccc ccccccgccc 6901 ccatcgctgc acaaaataat taaaaaataa ataaatacaa aattgggggt ggggaggggg 6961 gggagatggg gagagtgaag cagaacgtgg ggctcacctc gaccatggta atagcgatga 7021 ctaatacgta gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat 7081 gccaggcggg ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca 7141 cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga 7201 aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt 7261 cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat 7321 atgggctatg aactaatgac cccgtaattg attactatta ataactagtc aataatcaat 7381 gtcgagaccc atagagccca ccgcatcccc agcatgcctg ctattgtctt cccaatcctc 7441 ccccttgctg tcctgcccca ccccaccccc cagaatagaa tgacacctac tcagacaatg 7501 cgatgcaatt tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa 7561 ggaaggcacg ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc 7621 tgatcagcgg gtttataact tcgtataatg tatgctatac gaagttatgg taccttactc 7681 gtccatgccg ccggtggagt ggcggccctc ggcgcgttcg tactgttcca cgaatttaaa 7741 ttcaggcgcc aggctttctg gtcatgcacc aggttctagg gccctcaggc acttccacgt 7801 cggcggtcac ggtgaagccc agtctctcgt agaagggcag gtttctgggg gcgcttgttt 7861 ccaggaaggc gggcacgcca gccctttcag cagcttccac gccaggcagc accacagcag 7921 atcccagtcc cttgccctgg tggtcaggtg acacgcccac ggtggccaga aaccaggcag 7981 gctctttagg tctgtggggg gccagcaggc cttccatctg ctgctgggca gccagtctag 8041 agccgctcag ctcggccatt ctaggtccga tctcggcgaa cacagcgccg gcttccacag 8101 actcaggggt tgtccacaca gccacagcgg cgccatcatc ggccacccac actttgccga 8161 tgtccaggcc cactctggtc agaaacagtt cctgcagctc ggtcactctc tcgatgtgtc 8221 tgtcggggtc cacggtgtgt ctggtggcag ggtaatcggc gaaggcagcg gccagtgtcc 8281 gcacagctct aggcacatcg tccctggtgg ccagccgcac tgtgggcttg tactcggtca 8341 tagggccggg attctcctcc acgtcaccgc atgttagaag acttcctctg ccctcgccag 8401 agcctctgct gtgcatcttg tacagctcgt ccatgccgag agtgatcccg gcggcggtca 8461 cgaactccag caggaccatg tgatcgcgct tctcgttggg gtctttgctc agggcggact 8521 gggtgctcag gtagtggttg tcgggcagca gcacggggcc gtcgccgatg ggggtgttct 8581 gctggtagtg gtcggcgagc tgcacgctgc cgtcctcgat gttgtggcgg atcttgaagt 8641 tcaccttgat gccgttcttc tgcttgtcgg ccatgatata gacgttgtgg ctgttgtagt 8701 tgtactccag cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg cccttcagct 8761 cgatgcggtt caccagggtg tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc 8821 cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga 8881 agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa gcactgcacg ccgtaggtca 8941 gggtggtcac gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag atgaacttca 9001 gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc ggacacgctg aacttgtggc 9061 cgtttacgtc gccgtccagc tcgaccagga tgggcaccac cccggtgaac agctcctcgc 9121 ccttgctcac catggtggcg ctagcgtagg cgccggtcac agcttggatc tgtaacggcg 9181 cagaacagaa aacgaaacaa agacgtagag ttgagcaagc agggtcaggc aaagcgtgga 9241 gagccggctg agtctaggta ggctccaagg gagcgccgga caaaggcccg gtctcgacct 9301 gagctttaaa cttacctaga cggcggacgc agttcaggag gcaccacagg cgggaggcgg 9361 cagaacgcga ctcaaccggc gtggatggcg gcctcaggta gggcggcggg cgcgtgaagg 9421 agagatgcga gcccctcgaa gcttcagctg tgttctggcg gcaaacccgt tgcgaaaaag 9481 aacgttcacg gcgactactg cacttatata cggttctccc ccaccctcgg gaaaaaggcg 9541 gagccagtac acgacatcac tttcccagtt taccccgcgc caccttctct aggcaccggt 9601 tcaattgccg acccctcccc ccaacttctc ggggactgtg ggcgatgtgc gctctgccca 9661 ctgacgggca ccggagcgat cgcagatcga gagagaaatg ttctggcacc tgcacttgca 9721 ctggggacag cctattttgc tagtttgttt tgtttcgttt tgttttgatg gagagcgtat 9781 gttagtacta tcgattcaca caaaaaacca acacacagat gtaatgaaaa taaagatatt 9841 ttatttcacc gcttgaaggc gatgtatgcc accagtccga cgacgacggc tgccaggatg 9901 gagcaataca cgggaatcag gttatctgtg gtgccccggg tgaccactgg ctgggagctc 9961 cccatgacag ttgtgaccac gccagccacg gtggaggcga tcagatcctg ctctggtggt 10021 gcttctggct cctgtgtaga tggtgcggtg ctgtctgatc cttcaggggg ggtggaccga 10081 gtaatccatc gccctgggat ttcctcgcac tcggcgtctg cccatctggt acattcgcgc 10141 agctgccttt cggtatcctc gcacacagta caaggcaggc aggggtccac gtgattggcc 10201 tcatcgctat atgtcccgtc aggacattcc tcgcacacgg tgttctgttt atcctggcag 10261 ctaaagacca gccctgagcc tgcctcacac accctgcaag cttcacatct gccggtagtc 10321 tcgtcctgat agtatccgta tgcacaccgg cagactgcat cgtcagcctc cacacaagga 10381 gcactcatag actgcaggcc cacgcactca gtacatggct tgcaaggttc tgtggcactg 10441 accacatcag agaatgtgac ggagtccaga caaggctcgc acacagtctg attggctccg 10501 catggctgtg ccactccctc tcccaggtta caagctttac agcattctcc agaatgagtg 10561 tacagccctg tggggcaggc ttccttcgcg atcgctccac ttcctggctc tcgctgtccc 10621 tgaggatggt gttccagtct agcccgcagc tgttccagct ctcctgccag tccctcgacc 10681 acgtgtcctc ctcgccatcc tgtggtcttt tcgtccatcc ctgacccctc gagctgcacc 10741 tgtgaatgca ctccagttgc ggttgcgacc agaaacagaa tgatacatga ccagcccata 10801 gggccgggat tctcctccac gtcaccgcat gttagaagac ttcctctgcc ctctccgctg 10861 ccagatctct cgaggccctg tgggaggaag agaagaggtc agataacttc gtataatgta 10921 tgctatacga agttatgcgg ccgctgggag aagggtaaga gagctcaatc ctcatcttac 10981 atagaaggaa gtcagtagat aatatcttac atttaaaagt caagaaataa cagtctgacc 11041 atactatcta gaaatatgga agttaacagt aaggaagtta aaacagttga aagtagtttc 11101 gtctagaaag aaagacttgg ggaagaacaa agaaggctaa ttttcattat gagctttatt 11161 gtaacagaaa agagaccact aactcagcta ggttttccca attcctcaac actttggggc 11221 acagtgttta gtcagagctg tgatcactct ctgagcacat ggattgtatc ttaattttca 11281 tctcattgaa aatatgtacc atttgtccaa gtggaaactg ggctgaagca tgactgtgac 11341 caccaccaaa gaaatggcta gactcgctaa ggtctgctgg ggtgacccgt gattgtgttg 11401 tgcttgatct tgttgagcat ggctctttta agtcaccata cctgcccctg ccaatgcacc 11461 aaacaccaac ttaaagggac tttcagcttt tgtaagtcag tctcagcctt acttctgaac 11521 catttctttg acttctttca tgactctgat cctgtcccag ttttaagcct cccagccacc 11581 agtcatgctg ttttgctctg ctatttacat tcttctctac ctactttgga cttgatggta 11641 tgcctactat attagttatt tattgctgtg taacaaacca ctctgaaaca tagtggctta 11701 caaccacaac cattttattt tattatgaaa ctgtgggtca gaatgtatat tggcccattg 11761 gcatgattct cctgctattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc 11821 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg 11881 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc 11941 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact 12001 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc 12061 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag 12121 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat 12181 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt 12241 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa 12301 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc 12361 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg 12421 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt 12481 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca 12541 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat 12601 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca 12661 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg 12721 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg 12781 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt 12841 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg 12901 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata 12961 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca 13021 ccgcctacat acctcgctct gctaatcc //