出品公司: | Clontech |
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载体名称: | pRetroQ-DsRed-Monomer-N1 |
质粒类型: | 逆病毒载体;荧光报告载体 |
高拷贝/低拷贝: | 低拷贝 |
克隆方法: | 限制性内切酶,多克隆位点 |
启动子: | CMV IE |
载体大小: | 7571 bp |
5' 测序引物及序列: | -- |
3' 测序引物及序列: | -- |
载体标签: | DsRed-Monomer(C-端) |
载体抗性: | 氨苄青霉素 |
筛选标记: | 嘌呤霉素(Puromycin) |
克隆菌株: | DH5α |
宿主细胞(系): | 常规细胞系(293、CV-1、CHO等),原代细胞 |
备注: |
pRetroQ-DsRed-Monomer-N1载体是自我失活型逆病毒载体,表达C端DsRed-Monomer融合蛋白; pRetroQ-DsRed-Monomer-N1载体能够高效递送目的基因到靶细胞中,尤其适用于原代细胞和其它难转染的细胞系; DsRed-Monomer是DsRed的单体突变体,与DsRed相比,编码序列经过了人工优化,适用于在哺乳动物细胞中的高水平表达。 |
产品目录号: | 632507 |
稳定性: | 稳表达 |
组成型/诱导型: | 组成型 |
病毒/非病毒: | 逆转录病毒 |
买家导航
pRetroQ-DsRed-Monomer-N1载体质粒基本信息
pRetroQ-DsRed-Monomer-N1质粒图谱载体图谱和pRetroQ-DsRed-Monomer-N1载体序列质粒序列多克隆位点信息
pRetroQ-DsRed-Monomer-N1质粒载体简介
pRetroQ-DsRed Monomer-N1载体描述
pRetroQ-DsRed Monomer-N1 is a high-titer, self-inactivating retroviral vector that facilitates efficient delivery and expression of DsRed-monomer (DsRed.M1) as well as N terminal fusions of DsRed monomer to target cells. DsRed.M1 is a monomeric mutant derived from the tetrameric Discosoma sp. red fluorescent protein DsRed (1). DsRed-Monomer contains forty-five amino acid substitutions. When DsRed-Monomer is expressed in mammalian cell cultures, red fluorescent cells can be detected by either fluorescence microscopy or flow cytometry 12–16 hours after transfection or 24–48 hours after infection (DsRed-Monomer excitation and emission maxima = 557 nm and 592 nm, respectively. The DsRed-Monomer coding sequence is human codon-optimized for high expression in mammalian cells (2) The MCS in pRetroQ-DsRed Monomer-N1 lies between the immediate early promoter of CMV (PCMV IE) and the DsRed-Monomer coding sequences. Genes cloned into the MCS are expressed as fusions to the N-terminus of DsRed Monomer when they are in the same reading frame as DsRed Monomer and there are no intervening stop codons. The RetroQ retroviral vector backbone incorporates several unique features. This vector contains a puromycin resistance cassette (Puror) driven by the PGK promoter for selection of positively-infected cells (2).The hybrid 5’ long terminal repeat (LTR) consists of the CMV type I enhancer and the murine sarcoma virus (MSV) promoter. This vector demonstrates high levels of transcription in HEK 293-based packaging cell lines due, in part, to the presence of adenoviral E1A (3–6) in these cells. The self-inactivating feature of the vector is provided by a deletion in the 3’ LTR enhancer region (U3). During reverse transcription of the retroviral RNA, the inactivated 3’ LTR is copied and replaces the 5’ LTR CMV enhancer sequences. This can reduce the phenomenon known as promoter interference (7) and allow more efficient expression.
Additionally, the viral genomic transcript contains the necessary viral RNA processing elements, including the LTRs, packaging signal (ψ+), and tRNA primer-binding site. pRetroQDsRed Monomer-N1 contains a bacterial origin of replication, an E. coli Ampr gene for propagation and selection in bacteria, and an SV40 origin for replication in mammalian cells expressing the SV40 T antigen.
pRetroQ-DsRed Monomer-N1 is designed to efficiently deliver and express fusions to the N terminus of DsRed-Monomer into primary cells or cells that are difficult to transfect. Fusions to the N terminus of DsRed-Monomer retain the fluorescent properties of the native protein, allowing the in vivo localization of the fusion protein. The target gene should be cloned into pDsRed Monomer-N1 so that it is in frame with the DsRed-Monomer coding sequences, with no intervening in-frame stop codons. The inserted gene should include the initiating ATG codon. The recombinant DsRed Monomer vector can be infected or transfected into mammalian cells. If required, stable transformants can be selected using puromycin. pRetroQ DsRed Monomer-N1 can also be used simply to express DsRed Monomer in a cell line of interest (e.g., as an infection marker). Once pRetroQ-DsRed Monomer-N1 is transfected into a packaging cell line (such as the RetroPack PT67 Cell line, AmphoPack-293, EcoPack2-293, Pantropic Expression System , or Retro-X Universal Packaging System, RNA from the vector is packaged into non-infectious, replication-incompetent retroviral particles, since pRetroQ-DsRed Monomer-N1 lacks the structural genes (gag, pol, and env) necessary for particle formation and replication. These genes, however, are stably integrated as part of the packaging cell genome. Once a high-titer supernatant is produced, these retroviral particles can infect target cells and transmit the gene of interest but cannot replicate within these cells due to the absence of viral structural genes. The separate introduction and integration of the structural genes into the packaging cell line minimizes the chances of producing replicationcompetent virus due to recombination events during cell proliferation.
Propagation in E. coli
Suitable host strains: DH5α, Fusion Blue, and other general purpose strains.
Selectable marker: plasmid confers resistance to ampicillin (100 μg/ml) in E. coli hosts.
E. coli replication origin: ColE1
Copy number: low
pRetroQ-DsRed-Monomer-N1质粒序列载体序列
LOCUS pRetroQ-DsRed-Monomer-N1 7571 bp DNA SYN DEFINITION pRetroQ-DsRed-Monomer-N1 ACCESSION KEYWORDS SOURCE ORGANISM other sequences; artificial sequences; vectors. FEATURES Location/Qualifiers source 1..7571 /organism="pRetroQ-DsRed-Monomer-N1" /mol_type="other DNA" promoter 2..506 /label="CMV_immearly_promoter" misc_feature 6..688 /label="5_LTR" misc_feature 9..296 /label="CAG_enhancer" misc_feature 463..483 /label="CMV_fwd_primer" misc_feature 758..1567 /label="psi_plus_pack" misc_feature 1154..1561 /label="gag" misc_feature 1497..1519 /label="MSCV_primer" misc_feature 1497..1519 /label="pLXSN_5_primer" misc_feature 1535..1551 /label="pBABE_5_primer" promoter 1591..2143 /label="CMV_immearly_promoter" misc_feature 1646..1933 /label="CAG_enhancer" misc_feature 2100..2120 /label="CMV_fwd_primer" promoter 2101..2170 /label="CMV_promoter" CDS 2260..2937 /label="ORF frame 1" /translation="MDNTEDVIKEFMQFKVRMEGSVNGHYFEIEGEGEGKPYEGTQTA KLQVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYMKLSFPEGFTWERSMNFEDG GVVEVQQDSSLQDGTFIYKVKFKGVNFPADGPVMQKKTAGWEPSTEKLYPQDGVLKGE ISHALKLKDGGHYTCDFKTVYKAKKPVQLPGNHYVDSKLDITNHNEDYTVVEQYEHAE ARHSGSQ*" gene 2263..2934 /label="DsRed_Monomer" /gene="DsRed_Monomer" /translation="MDNTEDVIKEFMQFKVRMEGSVNGHYFEIEGEGEGKPYEGTQTA KLQVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYMKLSFPEGFTWERSMNFEDG GVVEVQQDSSLQDGTFIYKVKFKGVNFPADGPVMQKKTAGWEPSTEKLYPQDGVLKGE ISHALKLKDGGHYTCDFKTVYKAKKPVQLPGNHYVDSKLDITNHNEDYTVVEQYEHAE ARHSGSQ*" misc_feature complement(2440..2460) /label="dsRed1_N_primer" /translation="MDNTEDVIKEFMQFKVRMEGSVNGHYFEIEGEGEGKPYEGTQTA KLQVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYMKLSFPEGFTWERSMNFEDG GVVEVQQDSSLQDGTFIYKVKFKGVNFPADGPVMQKKTAGWEPSTEKLYPQDGVLKGE ISHALKLKDGGHYTCDFKTVYKAKKPVQLPGNHYVDSKLDITNHNEDYTVVEQYEHAE ARHSGSQ*" misc_feature complement(3001..3024) /label="MSCV_rev_primer" /translation="MDNTEDVIKEFMQFKVRMEGSVNGHYFEIEGEGEGKPYEGTQTA KLQVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYMKLSFPEGFTWERSMNFEDG GVVEVQQDSSLQDGTFIYKVKFKGVNFPADGPVMQKKTAGWEPSTEKLYPQDGVLKGE ISHALKLKDGGHYTCDFKTVYKAKKPVQLPGNHYVDSKLDITNHNEDYTVVEQYEHAE ARHSGSQ*" CDS 3246..4085 /label="ORF frame 3" /translation="MEAGRPLGQRPIAALLLRFLGSEAGKGWVRGRAQGRAQGRGGRP KVLRRPGILHASKAHVCRAVLLFLISGPFDLQPKLTMTEYKPTVRLATRDDVPRAVRT LAAAFADYPATRHTVDPDRHIERVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPE SVEAGAVFAEIGPRMAELSGSRLAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKG LGSAVVLPGVEAAERAGVPAFLETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRK PGA*" misc_feature 3398..3417 /label="mPGK_F_primer" /translation="MEAGRPLGQRPIAALLLRFLGSEAGKGWVRGRAQGRAQGRGGRP KVLRRPGILHASKAHVCRAVLLFLISGPFDLQPKLTMTEYKPTVRLATRDDVPRAVRT LAAAFADYPATRHTVDPDRHIERVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPE SVEAGAVFAEIGPRMAELSGSRLAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKG LGSAVVLPGVEAAERAGVPAFLETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRK PGA*" CDS complement(3411..4136) /label="ORF frame 1" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" gene 3486..4085 /label="puro (variant)" /gene="puro (variant)" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" misc_feature complement(4805..4827) /label="pGEX_3_primer" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" misc_feature complement(4963..4983) /label="pBABE_3_primer" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" misc_feature complement(4969..5184) /label="SV40_enhancer" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" promoter 4981..5249 /label="SV40_promoter" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" rep_origin 5148..5225 /label="SV40_origin" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" misc_feature 5210..5229 /label="SV40pro_F_primer" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" gene complement(6349..7209) /label="Ampicillin" /gene="Ampicillin" /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG KLGLQVERPGDEEEENSAADVRF*" CDS complement(6349..7209) /label="ORF frame 3" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW*" promoter complement(7251..7279) /label="AmpR_promoter" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW*" promoter 7501..506 /label="CMV_immearly_promoter" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW*" misc_feature 7565..688 /label="5_LTR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW*" ORIGIN 1 TCCGCGTTAC ATAACTTACG GTAAATGGCC CGCCTGGCTG ACCGCCCAAC GACCCCCGCC 61 CATTGACGTC AATAATGACG TATGTTCCCA TAGTAACGCC AATAGGGACT TTCCATTGAC 121 GTCAATGGGT GGAGTATTTA CGGTAAACTG CCCACTTGGC AGTACATCAA GTGTATCATA 181 TGCCAAGTAC GCCCCCTATT GACGTCAATG ACGGTAAATG GCCCGCCTGG CATTATGCCC 241 AGTACATGAC CTTATGGGAC TTTCCTACTT GGCAGTACAT CTACGTATTA GTCATCGCTA 301 TTACCATGGT GATGCGGTTT TGGCAGTACA TCAATGGGCG TGGATAGCGG TTTGACTCAC 361 GGGGATTTCC AAGTCTCCAC CCCATTGACG TCAATGGGAG TTTGTTTTGG CACCAAAATC 421 AACGGGACTT TCCAAAATGT CGTAACAACT CCGCCCCATT GACGCAAATG GGCGGTAGGC 481 GTGTACGGTG GGAGGTCTAT ATAAGCAGAG CTCAATAAAA GAGCCCACAA CCCCTCACTC 541 GGCGCGCCAG TCTTCCGATA GACTGCGTCG CCCGGGTACC CGTATTCCCA ATAAAGCCTC 601 TTGCTGTTTG CATCCGAATC GTGGTCTCGC TGTTCCTTGG GAGGGTCTCC TCTGAGTGAT 661 TGACTACCCA CGACGGGGGT CTTTCATTTG GGGGCTCGTC CGGGATTTGG AGACCCCTGC 721 CCAGGGACCA CCGACCCACC ACCGGGAGGT AAGCTGGCCA GCAACTTATC TGTGTCTGTC 781 CGATTGTCTA GTGTCTATGT TTGATGTTAT GCGCCTGCGT CTGTACTAGT TAGCTAACTA 841 GCTCTGTATC TGGCGGACCC GTGGTGGAAC TGACGAGTTC TGAACACCCG GCCGCAACCC 901 TGGGAGACGT CCCAGGGACT TTGGGGGCCG TTTTTGTGGC CCGACCTGAG GAAGGGAGTC 961 GATGTGGAAT CCGACCCCGT CAGGATATGT GGTTCTGGTA GGAGACGAGA ACCTAAAACA 1021 GTTCCCGCCT CCGTCTGAAT TTTTGCTTTC GGTTTGGAAC CGAAGCCGCG CGTCTTGTCT 1081 GCTGCAGCGC TGCAGCATCG TTCTGTGTTG TCTCTGTCTG ACTGTGTTTC TGTATTTGTC 1141 TGAAAATTAG GGCCAGACTG TTACCACTCC CTTAAGTTTG ACCTTAGGTC ACTGGAAAGA 1201 TGTCGAGCGG ATCGCTCACA ACCAGTCGGT AGATGTCAAG AAGAGACGTT GGGTTACCTT 1261 CTGCTCTGCA GAATGGCCAA CCTTTAACGT CGGATGGCCG CGAGACGGCA CCTTTAACCG 1321 AGACCTCATC ACCCAGGTTA AGATCAAGGT CTTTTCACCT GGCCCGCATG GACACCCAGA 1381 CCAGGTCCCC TACATCGTGA CCTGGGAAGC CTTGGCTTTT GACCCCCCTC CCTGGGTCAA 1441 GCCCTTTGTA CACCCTAAGC CTCCGCCTCC TCTTCCTCCA TCCGCCCCGT CTCTCCCCCT 1501 TGAACCTCCT CGTTCGACCC CGCCTCGATC CTCCCTTTAT CCAGCCCTCA CTCCTTCTCT 1561 AGGCGCCGGA ATTGAAGATC ATAGTTATTA ATAGTAATCA ATTACGGGGT CATTAGTTCA 1621 TAGCCCATAT ATGGAGTTCC GCGTTACATA ACTTACGGTA AATGGCCCGC CTGGCTGACC 1681 GCCCAACGAC CCCCGCCCAT TGACGTCAAT AATGACGTAT GTTCCCATAG TAACGCCAAT 1741 AGGGACTTTC CATTGACGTC AATGGGTGGA GTATTTACGG TAAACTGCCC ACTTGGCAGT 1801 ACATCAAGTG TATCATATGC CAAGTACGCC CCCTATTGAC GTCAATGACG GTAAATGGCC 1861 CGCCTGGCAT TATGCCCAGT ACATGACCTT ATGGGACTTT CCTACTTGGC AGTACATCTA 1921 CGTATTAGTC ATCGCTATTA CCATGGTGAT GCGGTTTTGG CAGTACATCA ATGGGCGTGG 1981 ATAGCGGTTT GACTCACGGG GATTTCCAAG TCTCCACCCC ATTGACGTCA ATGGGAGTTT 2041 GTTTTGGCAC CAAAATCAAC GGGACTTTCC AAAATGTCGT AACAACTCCG CCCCATTGAC 2101 GCAAATGGGC GGTAGGCGTG TACGGTGGGA GGTCTATATA AGCAGAGCTG GTTTAGTGAA 2161 CCGTCAGATC CGCTAGCGCT ACCGGACTCA GATCTCGAGC TCAAGCTTCG AATTCTGCAG 2221 TCGACGGTAC CGCGGGCCCG GGATCCACCG GTCGCCACCA TGGACAACAC CGAGGACGTC 2281 ATCAAGGAGT TCATGCAGTT CAAGGTGCGC ATGGAGGGCT CCGTGAACGG CCACTACTTC 2341 GAGATCGAGG GCGAGGGCGA GGGCAAGCCC TACGAGGGCA CCCAGACCGC CAAGCTGCAG 2401 GTGACCAAGG GCGGCCCCCT GCCCTTCGCC TGGGACATCC TGTCCCCCCA GTTCCAGTAC 2461 GGCTCCAAGG CCTACGTGAA GCACCCCGCC GACATCCCCG ACTACATGAA GCTGTCCTTC 2521 CCCGAGGGCT TCACCTGGGA GCGCTCCATG AACTTCGAGG ACGGCGGCGT GGTGGAGGTG 2581 CAGCAGGACT CCTCCCTGCA GGACGGCACC TTCATCTACA AGGTGAAGTT CAAGGGCGTG 2641 AACTTCCCCG CCGACGGCCC CGTAATGCAG AAGAAGACTG CCGGCTGGGA GCCCTCCACC 2701 GAGAAGCTGT ACCCCCAGGA CGGCGTGCTG AAGGGCGAGA TCTCCCACGC CCTGAAGCTG 2761 AAGGACGGCG GCCACTACAC CTGCGACTTC AAGACCGTGT ACAAGGCCAA GAAGCCCGTG 2821 CAGCTGCCCG GCAACCACTA CGTGGACTCC AAGCTGGACA TCACCAACCA CAACGAGGAC 2881 TACACCGTGG TGGAGCAGTA CGAGCACGCC GAGGCCCGCC ACTCCGGCTC CCAGTAGAGC 2941 GGCCGCGACT CTAGATAATT CTACCGGGTA GGGGAGGCGC TTTTCCCAAG GCAGTCTGGA 3001 GCATGCGCTT TAGCAGCCCC GCTGGGCACT TGGCGCTACA CAAGTGGCCT CTGGCCTCGC 3061 ACACATTCCA CATCCACCGG TAGGCGCCAA CCGGCTCCGT TCTTTGGTGG CCCCTTCGCG 3121 CCACCTTCTA CTCCTCCCCT AGTCAGGAAG TTCCCCCCCG CCCCGCAGCT CGCGTCGTGC 3181 AGGACGTGAC AAATGGAAGT AGCACGTCTC ACTAGTCTCG TGCAGATGGA CAGCACCGCT 3241 GAGCAATGGA AGCGGGTAGG CCTTTGGGGC AGCGGCCAAT AGCAGCTTTG CTCCTTCGCT 3301 TTCTGGGCTC AGAGGCTGGG AAGGGGTGGG TCCGGGGGCG GGCTCAGGGG CGGGCTCAGG 3361 GGCGGGGCGG GCGCCCGAAG GTCCTCCGGA GGCCCGGCAT TCTGCACGCT TCAAAAGCGC 3421 ACGTCTGCCG CGCTGTTCTC CTCTTCCTCA TCTCCGGGCC TTTCGACCTG CAGCCCAAGC 3481 TTACCATGAC CGAGTACAAG CCCACGGTGC GCCTCGCCAC CCGCGACGAC GTCCCCAGGG 3541 CCGTACGCAC CCTCGCCGCC GCGTTCGCCG ACTACCCCGC CACGCGCCAC ACCGTCGATC 3601 CGGACCGCCA CATCGAGCGG GTCACCGAGC TGCAAGAACT CTTCCTCACG CGCGTCGGGC 3661 TCGACATCGG CAAGGTGTGG GTCGCGGACG ACGGCGCCGC GGTGGCGGTC TGGACCACGC 3721 CGGAGAGCGT CGAAGCGGGG GCGGTGTTCG CCGAGATCGG CCCGCGCATG GCCGAGTTGA 3781 GCGGTTCCCG GCTGGCCGCG CAGCAACAGA TGGAAGGCCT CCTGGCGCCG CACCGGCCCA 3841 AGGAGCCCGC GTGGTTCCTG GCCACCGTCG GCGTCTCGCC CGACCACCAG GGCAAGGGTC 3901 TGGGCAGCGC CGTCGTGCTC CCCGGAGTGG AGGCGGCCGA GCGCGCCGGG GTGCCCGCCT 3961 TCCTGGAGAC CTCCGCGCCC CGCAACCTCC CCTTCTACGA GCGGCTCGGC TTCACCGTCA 4021 CCGCCGACGT CGAGGTGCCC GAAGGACCGC GCACCTGGTG CATGACCCGC AAGCCCGGTG 4081 CCTGACGCCC GCCCCACGAC CCGCAGCGCC CGACCGAAAG GAGCGCACGA CCCCATGCAT 4141 CGGCGTCTCG ATCGAGATAT CAGTGGTCCA GGCTCTAGTT TTGACTCAAC AATATCACCA 4201 GCTGAAGCCT ATAGAGTACG AGCCATAGAT AAAATAAAAG ATTTTATTTA GTCTCCAGAA 4261 AAAGGGGGGA ATGAAAGACC CCACCTGTAG GTTTGGCAAG CTACTAGCTT AAGTAACGCC 4321 ATTTTGCAAG GCATGGAAAA ATACATAACT GAGAATAGAG AAGTTCAGAT CAAGGTCAGG 4381 AACAGATGGA ACAGGGTCGA TCGACCCTAG AGAACCATCA GATGTTTCCA GGGTGCCCCA 4441 AGGACCTGAA ATGACCCTGT GCCTTATTTG AACTAACCAA TCAGTTCGCT TCTCGCTTCT 4501 GTTCGCGCGC TTCTGCTCCC CGAGCTCAAT AAAAGAGCCC ACAACCCCTC ACTCGGGGCG 4561 CCAGTCCTCC GATTGACTGA GTCGCCCGGG TACCCGTGTA TCCAATAAAC CCTCTTGCAG 4621 TTGCATCCGA CTTGTGGTCT CGCTGTTCCT TGGGAGGGTC TCCTCTGAGT GATTGACTAC 4681 CCGTCAGCGG GGGTCTTTCA TTTGGGGGCT CGTCCGGGAT CGGGAGACCC CTGCCCAGGG 4741 ACCACCGACC CACCACCGGG AGGTAAGCTG GCTGCCTCGC GCGTTTCGGT GATGACGGTG 4801 AAAACCTCTG ACACATGCAG CTCCCGGAGA CGGTCACAGC TTGTCTGTAA GCGGATGCCG 4861 GGAGCAGACA AGCCCGTCAG GGCGCGTCAG CGGGTGTTGG CGGGTGTCGG GGCGCAGCCA 4921 TGACCCAGTC ACGTAGCGAT AGCGGAGTGT AGATCCGGCT GTGGAATGTG TGTCAGTTAG 4981 GGTGTGGAAA GTCCCCAGGC TCCCCAGCAG GCAGAAGTAT GCAAAGCATG CATCTCAATT 5041 AGTCAGCAAC CAGGTGTGGA AAGTCCCCAG GCTCCCCAGC AGGCAGAAGT ATGCAAAGCA 5101 TGCATCTCAA TTAGTCAGCA ACCATAGTCC CGCCCCTAAC TCCGCCCATC CCGCCCCTAA 5161 CTCCGCCCAG TTCCGCCCAT TCTCCGCCCC ATGGCTGACT AATTTTTTTT ATTTATGCAG 5221 AGGCCGAGGC CGCCTCGGCC TCTGAGCTAT TCCAGAAGTA GTGAGGAGGC TTTTTTGGAG 5281 GCCTAGGCTT TTGCAAAAAG CTTACTGGCT TAACTATGCG GCATCAGAGC AGATTGTACT 5341 GAGAGTGCAC CATATGCGGT GTGAAATACC GCACAGATGC GTAAGGAGAA AATACCGCAT 5401 CAGGCGCTCT TCCGCTTCCT CGCTCACTGA CTCGCTGCGC TCGGTCGTTC GGCTGCGGCG 5461 AGCGGTATCA GCTCACTCAA AGGCGGTAAT ACGGTTATCC ACAGAATCAG GGGATAACGC 5521 AGGAAAGAAC ATGTGAGCAA AAGGCCAGCA AAAGGCCAGG AACCGTAAAA AGGCCGCGTT 5581 GCTGGCGTTT TTCCATAGGC TCCGCCCCCC TGACGAGCAT CACAAAAATC GACGCTCAAG 5641 TCAGAGGTGG CGAAACCCGA CAGGACTATA AAGATACCAG GCGTTTCCCC CTGGAAGCTC 5701 CCTCGTGCGC TCTCCTGTTC CGACCCTGCC GCTTACCGGA TACCTGTCCG CCTTTCTCCC 5761 TTCGGGAAGC GTGGCGCTTT CTCATAGCTC ACGCTGTAGG TATCTCAGTT CGGTGTAGGT 5821 CGTTCGCTCC AAGCTGGGCT GTGTGCACGA ACCCCCCGTT CAGCCCGACC GCTGCGCCTT 5881 ATCCGGTAAC TATCGTCTTG AGTCCAACCC GGTAAGACAC GACTTATCGC CACTGGCAGC 5941 AGCCACTGGT AACAGGATTA GCAGAGCGAG GTATGTAGGC GGTGCTACAG AGTTCTTGAA 6001 GTGGTGGCCT AACTACGGCT ACACTAGAAG GACAGTATTT GGTATCTGCG CTCTGCTGAA 6061 GCCAGTTACC TTCGGAAAAA GAGTTGGTAG CTCTTGATCC GGCAAACAAA CCACCGCTGG 6121 TAGCGGTGGT TTTTTTGTTT GCAAGCAGCA GATTACGCGC AGAAAAAAAG GATCTCAAGA 6181 AGATCCTTTG ATCTTTTCTA CGGGGTCTGA CGCTCAGTGG AACGAAAACT CACGTTAAGG 6241 GATTTTGGTC ATGAGATTAT CAAAAAGGAT CTTCACCTAG ATCCTTTTAA ATTAAAAATG 6301 AAGTTTTAAA TCAATCTAAA GTATATATGA GTAAACTTGG TCTGACAGTT ACCAATGCTT 6361 AATCAGTGAG GCACCTATCT CAGCGATCTG TCTATTTCGT TCATCCATAG TTGCCTGACT 6421 CCCCGTCGTG TAGATAACTA CGATACGGGA GGGCTTACCA TCTGGCCCCA GTGCTGCAAT 6481 GATACCGCGA GACCCACGCT CACCGGCTCC AGATTTATCA GCAATAAACC AGCCAGCCGG 6541 AAGGGCCGAG CGCAGAAGTG GTCCTGCAAC TTTATCCGCC TCCATCCAGT CTATTAATTG 6601 TTGCCGGGAA GCTAGAGTAA GTAGTTCGCC AGTTAATAGT TTGCGCAACG TTGTTGCCAT 6661 TGCTGCAGGC ATCGTGGTGT CACGCTCGTC GTTTGGTATG GCTTCATTCA GCTCCGGTTC 6721 CCAACGATCA AGGCGAGTTA CATGATCCCC CATGTTGTGC AAAAAAGCGG TTAGCTCCTT 6781 CGGTCCTCCG ATCGTTGTCA GAAGTAAGTT GGCCGCAGTG TTATCACTCA TGGTTATGGC 6841 AGCACTGCAT AATTCTCTTA CTGTCATGCC ATCCGTAAGA TGCTTTTCTG TGACTGGTGA 6901 GTACTCAACC AAGTCATTCT GAGAATAGTG TATGCGGCGA CCGAGTTGCT CTTGCCCGGC 6961 GTCAACACGG GATAATACCG CGCCACATAG CAGAACTTTA AAAGTGCTCA TCATTGGAAA 7021 ACGTTCTTCG GGGCGAAAAC TCTCAAGGAT CTTACCGCTG TTGAGATCCA GTTCGATGTA 7081 ACCCACTCGT GCACCCAACT GATCTTCAGC ATCTTTTACT TTCACCAGCG TTTCTGGGTG 7141 AGCAAAAACA GGAAGGCAAA ATGCCGCAAA AAAGGGAATA AGGGCGACAC GGAAATGTTG 7201 AATACTCATA CTCTTCCTTT TTCAATATTA TTGAAGCATT TATCAGGGTT ATTGTCTCAT 7261 GAGCGGATAC ATATTTGAAT GTATTTAGAA AAATAAACAA ATAGGGGTTC CGCGCACATT 7321 TCCCCGAAAA GTGCCACCTG ACGTCTAAGA AACCATTATT ATCATGACAT TAACCTATAA 7381 AAATAGGCGT ATCACGAGGC CCTTTCGTCT TCAAGAATTA GCTTGGCCAT TGCATACGTT 7441 GTATCCATAT CATAATATGT ACATTTATAT TGGCTCATGT CCAACATTAC CGCCATGTTG 7501 ACATTGATTA TTGACTAGTT ATTAATAGTA ATCAATTACG GGGTCATTAG TTCATAGCCC 7561 ATATATGGAG T //