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片段分析法 SNP 基因分型
SNP 基因分型可鉴别人类基因组中常见 DNA 变异的单核苷酸多态性 (SNP)。SNP 已被证明是造成遗传性状、疾病易感性和药物治疗应答差异的原因。
对于从事疾病研究和疾病治疗的研究人员而言，SNP 基因分型变得尤为重要。大约每隔100到300个碱基就存在一个 SNP，可使用 RFLP、SSCP、测序等各种不同技术对其进行检测。
图1.SNaPshot® 标记化学法
图1.SNaPshot® 标记化学法依赖于单个碱基的延长和终止。SNaPshot® 多通路检测试剂盒使用单管反应来研究已知位置的 SNP。该化学法基于单个未标记寡核苷酸引物（或多个引物）的双脱氧法单碱基延伸。在荧光标记的 ddNTP 和 DNA 聚合酶都存在的情况下，每个引物均与互补模板结合。聚合酶使引物延伸一个核苷酸长度，在其3'端添加单个 ddNTP。荧光颜色读数将报告添加的是哪个碱基。
SNaPshot® 多通路检测系统是一种基于引物延伸的 SNP 分析方法。其具有多通路检测能力，可在单次反应中分析多达10个 SNP，无论该 SNP 在染色体的哪个位置，也无论该 SNP 与相邻 SNP 位点的分离量有多大。能够使用未标记的用户自定义引物，从而研究人员可以较低成本自行掺入感兴趣的 SNP。多通路检测即用型反应混合物（包含在系统中）有助于确保对联合样品进行稳定、可重现的分析。研究人员只需一台3730xl基因分析仪，就可以每天分析23,000多个 SNP 基因分型。
SNaPshot® 多通路检测系统也可执行 BAC 指纹图谱分析和甲基化分析。
我们的客户使用 SNaPshot® 多通路检测系统通过多种方法进行他们的研究实践，请转至下方的“出版物和文献”选项卡，了解其中的部分方法。
SNP 基因分型逐步指南
 DNA 提取是进行 DNA 测序和片段分析实验流程中关键的第一步。样品本身特性以及所选核酸提取方法会对 DNA 测序读段的总体质量、准确性和长度产生明显影响。来源或组织类型、获取方式以及提取前样品处理或储存方式不同，理想方法也会不同。 推荐产品：DNA 分离 |
执行：SNaPshot 反应之前，必须去除引物和未掺入的 dNTP。 纯化：SNaPshot 反应后，使用牛小肠碱性磷酸酶对产品进行短暂处理十分重要。 推荐产品：执行和纯化 SNaPshot® 反应
| ||||||||||
将一份 SNaPshot 反应液的等分试样加入到由 Hi-Di 甲酰胺和 GeneScan LIZ-120 大小标准品组成的电泳溶液中。 推荐产品：制备待分析样品 |
 确定 SNP 基因分型 推荐产品：数据分析 |
DNA 提取是进行 DNA 测序和片段分析实验流程中关键的第一步。样品本身特性以及所选核酸提取方法会对 DNA 测序读段的总体质量、准确性和长度产生明显影响。来源或组织类型、获取方式以及提取前样品处理或储存方式不同，理想方法也会不同。 推荐产品：DNA 分离 |
执行：SNaPshot 反应之前，必须去除引物和未掺入的 dNTP。 纯化：SNaPshot 反应后，使用牛小肠碱性磷酸酶对产品进行短暂处理十分重要。 推荐产品：执行和纯化 SNaPshot® 反应
| ||||||||||
将一份 SNaPshot 反应液的等分试样加入到由 Hi-Di 甲酰胺和 GeneScan LIZ-120 大小标准品组成的电泳溶液中。 推荐产品：制备待分析样品 |
确定 SNP 基因分型 推荐产品：数据分析 |
产品文献
- 实验方案：ABI PRISM® SNaPshot® 多通路检测系统
- 快速参考卡：ABI PRISM® SNaPshot® 多通路检测系统
- 使用说明：在3730/3730xl DNA 分析仪和3130/3130xl基因分析仪上使用 SNaPshot®
- 应用资料：在 Applied Biosystems 3730/3730xl DNA 分析仪上进行 BAC 指纹图谱分析 (3 MB)
- 入门指南：GeneMapper® 软件4.0版和 SNaPshot® 试剂盒分析 (3.71 MB)
参考文献
SNP 分析
- 植物乳杆菌中的物种鉴定
Rapid identification of Lactobacillus plantarum group using the SNaPshot minisequencing assay.
Publication: Syst Appl Microbiol.2011 Dec;34(8):586-9.
Authors: Huang CH, Chang MT, Huang MC, Lee FL. - 老虎亚种鉴定
The development and validation of a single SNaPshot multiplex for tiger species and subspecies identification-Implications for forensic purposes.
Publication: Forensic Sci Int Genet.2011 Jun 30.
Authors: Kitpipit T, Tobe SS, Kitchener AC, Gill P, Linacre A. - 跟踪奶牛及其产品
SNPmplexViewer–toward a cost-effective traceability system.
Publication: BMC Res Notes.2011 May 23;4:146.
Authors: Seroussi Y, Shirak A, Seroussi E. - 鉴定：区分毒品大麻和非毒品大麻
Differentiation of drug and non-drug Cannabis using a single nucleotide polymorphism (SNP) assay.
Publication: Forensic Sci Int. 2011 Apr 15;207(1-3):193-7.Epub 2010 Oct 30.
Authors: Rotherham D, Harbison SA. - 鉴定结核分枝杆菌复合种中的物种
Identification and genotyping of Mycobacterium tuberculosis complex species by use of a SNaPshot Minisequencing-based assay.
Publication: J Clin Microbiol.2010 May;48(5):1758-66.Epub 2010 Mar 10.
Authors: Bouakaze C, Keyser C, de Martino SJ, Sougakoff W, Veziris N, Dabernat H, Ludes B. - 血型检测
Detection of blood group genes using multiplex SNaPshot method.
Publication: Transfusion.2009 Apr;49(4):740-9.Epub 2009 Jan 2.
Authors: Palacajornsuk P, Halter C, Isakova V, Tarnawski M, Farmar J, Reid ME, Chaudhuri A. - 马匹选择性育种
Identification of horse chestnut coat color genotype using SNaPshot.
Publication: BMC Res Notes.2009 Dec 16;2:255.
Authors: Rendo F, Iriondo M, Manzano C, Estonba A. - 小麦育种
Insertion site-based polymorphism markers open new perspectives for genome saturation and marker-assisted selection in wheat.
Publication: Plant Biotechnol J. 2010 Feb;8(2):196-210.
Authors: Paux E, Faure S, Choulet F, Roger D, Gauthier V, Martinant JP, Sourdille P, Balfourier F, Le Paslier MC, Chauveau A, Cakir M, Gandon B, Feuillet C. - 线粒体 SNP 筛选
A new method for analysis of mitochondrial DNA point mutations and assess levels of heteroplasmy.
Publication: Biochem Biophys Res Commun (2006) 342(2): 387-393.
Authors: Cassandrini, D., M. G. Calevo, et al. - 干酪乳杆菌中的物种鉴定
Application of the SNaPshot minisequencing assay to species identification in the Lactobacillus casei group.
Publication: Mol Cell Probes.2011 Aug;25(4):153-7.
Authors: Huang CH, Chang MT, Huang MC, Lee FL. - 新一代测序后验证异质性
Detecting heteroplasmy from high-throughput sequencing of complete human mitochondrial DNA genomes.
Publication: The American Journal of Human Genetics 2010, 87(2), 237-249.
Authors: Li M, Schonberg A, Schaefer M, et al. - 线粒体变异体
Mitochondrial variants in schizophrenia, bipolar disorder, and major depressive disorder.
Publication: PLoS One (2009)4(3): e4913.
Authors: Rollins, B., M. V. Martin, et al. - SNP 筛选
A SNaPshot assay for the rapid and simple detection of four common hotspot codon mutations in the PIK3CA gene.
Publication: BMC Research Notes 2009, 2:66
Authors: Hurst, D C,Zuiverloon, TCM, Hafner, C, Zwarthoff, EC et al. - 杂合性缺失
Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors.
Publication: Clin Chem 54(11): 1844-1854 (2008)
Authors: Bujalkova, M., K. Zavodna, et al. - 逆转录病毒抗性突变
Pre-screening HIV-1 reverse transcriptase resistance mutations in subtype B patients using a novel multiplex primer extension assay.
Publication: Curr HIV Res (2009) 7(4): 398-409.
Authors: Jakobsen, M. R., A. Aggerholm, et al. - 评估猪的肉用性能
SNaPshot minisequencing and a panel of candidate genes for complex routine testing of meat performance traits in pigs.
Publication: Anim Biotechnol.2007;18(2):109-15.
Authors: Civánová K, Knoll A. - 降解的 DNA
Developing multiplexed SNP assays with special reference to degraded DNA templates.
Publication: Nature Protocols 2006;1(3):1370-8
Authors: Sanchez, JJ.& Endicott, P. - 病原抗性
Single nucleotide polymorphism and haplotypes in the IL10 region associated with HCV clearance
Publication: Genes Immun.2005 Jun;6(4):347-57.
Authors: Oleksyk TK, Thio CL, Truelove AL, Goedert JJ, Donfield SM, Kirk GD, Thomas DL, O'Brien SJ, Smith MW. - 线粒体 SNP 筛选
Mitochondrial DNA mutations in patients with postlingual, nonsyndromic hearing impairment
Publication: European Journal of Human Genetics (2005) 13, 26−33.
Authors: Jacobs HT, Hutchin TP, Käppi T, Gillies G, et al. - 痒病易感性筛查
Primer extension assay for prion protein genotype determination in sheep.
Publication: Molecular and Cellular Probes (2004), 18(1), 33-37.
Authors: Vaccari G, Conte M, Morelli L, Di Guardo G, et al. - 癌变过程中的基因改变
Role of COX-2, thromboxane A2 synthase, and prostaglandin I2 synthase in papillary thyroid carcinoma growth.
Publication: Mod Pathol.2005 Feb;18(2):221-7.
Authors: Kajita S, Ruebel KH, Casey MB, Nakamura N et al. - 拟南芥标记物
Establishment of a high-efficiency SNP-based framework marker set for Arabidopsis.
Publication: Plant J. 2003 Oct;36(1):122-40.
Authors: Törjék O, Berger D, Meyer RC, Müssig C, Schmid KJ, Rosleff Sörensen T, Weisshaar B, Mitchell-Olds T, Altmann T. - 免疫中的 SNP 验证
Single nucleotide polymorphisms and haplotypes in the IL10 region associated with HCV clearance.
Authors: Oleksyk TK, Thio CL, Truelove AL, Goedert JJ, et al.
SNP 定量
- 突变体定量测定
Putative precursor cancer cells in human colorectal cancer tissue.
Publication: Int J Clin Exp Pathol (2009)2(2): 154-162
Authors: Goranova, T. E., M. Ohue, et al. - 等位基因特异性基因表达
Cis-acting variation in the expression of a high proportion of gene in human brain.
Publication: Hum Genet.2003 Jul;113(2):149-53.Epub 2003 May 1.
Authors: Bray NJ, Buckland PR, Owen MJ, O'Donovan MC. - 等位基因特异性基因表达
Allelic variation in human gene expression
Publication: Curr Opin Oncol.2004 Jan;16(1):39-43.
Authors: Yan H, Zhou W. - SNP 等位基因频率定量测定
Universal, robust, highly quantitative SNP allele frequency measurement in DNA pools.
Publication: Hum Genet.2002 May;110(5):471-8.Epub 2002 Mar 23.
Authors: Norton N, Williams NM, Williams HJ, Spurlock G et al.
BAC 指纹图谱分析
- 高通量细菌人工染色体指纹图谱分析
High-throughput fingerprinting of bacterial artificial chromosomes using
Publication: Genomics 82 (2003) 378–389
Authors: Ming-Cheng Luo, Carolyn Thomas, Frank M. You, Joseph Hsiao
甲基化
- 检测甲基化状态
Capillary electrophoretic analysis of methylation status in CpG-rich regions by single-base extension of primers modified with N6-methoxy-2,6-diaminopurine
Publication: Analytical Biochemistry Volume 380, Issue 1, 1 September 2008, Pages 13-20.
Authors: Boyd, V.L. et al. - 甲基化状态
Single nucleotide extension technology for quantitative site-specific evaluation of metC/C in GC-rich regions.
Publication: Nucleic Acids Res (2005) 33(10): e95.
Authors: Kaminsky, Z. A., A. Assadzadeh, et al.
软件下载
- 目前无软件可供下载。
SNP 基因分型逐步指南
DNA 提取是进行 DNA 测序和片段分析实验流程中关键的第一步。样品本身特性以及所选核酸提取方法会对 DNA 测序读段的总体质量、准确性和长度产生明显影响。来源或组织类型、获取方式以及提取前样品处理或储存方式不同，理想方法也会不同。 推荐产品：DNA 分离 |
执行：SNaPshot 反应之前，必须去除引物和未掺入的 dNTP。 纯化：SNaPshot 反应后，使用牛小肠碱性磷酸酶对产品进行短暂处理十分重要。 推荐产品：执行和纯化 SNaPshot® 反应
| ||||||||||
将一份 SNaPshot 反应液的等分试样加入到由 Hi-Di 甲酰胺和 GeneScan LIZ-120 大小标准品组成的电泳溶液中。 推荐产品：制备待分析样品 |
确定 SNP 基因分型 推荐产品：数据分析 |
DNA 提取是进行 DNA 测序和片段分析实验流程中关键的第一步。样品本身特性以及所选核酸提取方法会对 DNA 测序读段的总体质量、准确性和长度产生明显影响。来源或组织类型、获取方式以及提取前样品处理或储存方式不同，理想方法也会不同。 推荐产品：DNA 分离 |
执行：SNaPshot 反应之前，必须去除引物和未掺入的 dNTP。 纯化：SNaPshot 反应后，使用牛小肠碱性磷酸酶对产品进行短暂处理十分重要。 推荐产品：执行和纯化 SNaPshot® 反应
| ||||||||||
将一份 SNaPshot 反应液的等分试样加入到由 Hi-Di 甲酰胺和 GeneScan LIZ-120 大小标准品组成的电泳溶液中。 推荐产品：制备待分析样品 |
确定 SNP 基因分型 推荐产品：数据分析 |
产品文献
- 实验方案：ABI PRISM® SNaPshot® 多通路检测系统
- 快速参考卡：ABI PRISM® SNaPshot® 多通路检测系统
- 使用说明：在3730/3730xl DNA 分析仪和3130/3130xl基因分析仪上使用 SNaPshot®
- 应用资料：在 Applied Biosystems 3730/3730xl DNA 分析仪上进行 BAC 指纹图谱分析 (3 MB)
- 入门指南：GeneMapper® 软件4.0版和 SNaPshot® 试剂盒分析 (3.71 MB)
参考文献
SNP 分析
- 植物乳杆菌中的物种鉴定
Rapid identification of Lactobacillus plantarum group using the SNaPshot minisequencing assay.
Publication: Syst Appl Microbiol.2011 Dec;34(8):586-9.
Authors: Huang CH, Chang MT, Huang MC, Lee FL. - 老虎亚种鉴定
The development and validation of a single SNaPshot multiplex for tiger species and subspecies identification-Implications for forensic purposes.
Publication: Forensic Sci Int Genet.2011 Jun 30.
Authors: Kitpipit T, Tobe SS, Kitchener AC, Gill P, Linacre A. - 跟踪奶牛及其产品
SNPmplexViewer–toward a cost-effective traceability system.
Publication: BMC Res Notes.2011 May 23;4:146.
Authors: Seroussi Y, Shirak A, Seroussi E. - 鉴定：区分毒品大麻和非毒品大麻
Differentiation of drug and non-drug Cannabis using a single nucleotide polymorphism (SNP) assay.
Publication: Forensic Sci Int. 2011 Apr 15;207(1-3):193-7.Epub 2010 Oct 30.
Authors: Rotherham D, Harbison SA. - 鉴定结核分枝杆菌复合种中的物种
Identification and genotyping of Mycobacterium tuberculosis complex species by use of a SNaPshot Minisequencing-based assay.
Publication: J Clin Microbiol.2010 May;48(5):1758-66.Epub 2010 Mar 10.
Authors: Bouakaze C, Keyser C, de Martino SJ, Sougakoff W, Veziris N, Dabernat H, Ludes B. - 血型检测
Detection of blood group genes using multiplex SNaPshot method.
Publication: Transfusion.2009 Apr;49(4):740-9.Epub 2009 Jan 2.
Authors: Palacajornsuk P, Halter C, Isakova V, Tarnawski M, Farmar J, Reid ME, Chaudhuri A. - 马匹选择性育种
Identification of horse chestnut coat color genotype using SNaPshot.
Publication: BMC Res Notes.2009 Dec 16;2:255.
Authors: Rendo F, Iriondo M, Manzano C, Estonba A. - 小麦育种
Insertion site-based polymorphism markers open new perspectives for genome saturation and marker-assisted selection in wheat.
Publication: Plant Biotechnol J. 2010 Feb;8(2):196-210.
Authors: Paux E, Faure S, Choulet F, Roger D, Gauthier V, Martinant JP, Sourdille P, Balfourier F, Le Paslier MC, Chauveau A, Cakir M, Gandon B, Feuillet C. - 线粒体 SNP 筛选
A new method for analysis of mitochondrial DNA point mutations and assess levels of heteroplasmy.
Publication: Biochem Biophys Res Commun (2006) 342(2): 387-393.
Authors: Cassandrini, D., M. G. Calevo, et al. - 干酪乳杆菌中的物种鉴定
Application of the SNaPshot minisequencing assay to species identification in the Lactobacillus casei group.
Publication: Mol Cell Probes.2011 Aug;25(4):153-7.
Authors: Huang CH, Chang MT, Huang MC, Lee FL. - 新一代测序后验证异质性
Detecting heteroplasmy from high-throughput sequencing of complete human mitochondrial DNA genomes.
Publication: The American Journal of Human Genetics 2010, 87(2), 237-249.
Authors: Li M, Schonberg A, Schaefer M, et al. - 线粒体变异体
Mitochondrial variants in schizophrenia, bipolar disorder, and major depressive disorder.
Publication: PLoS One (2009)4(3): e4913.
Authors: Rollins, B., M. V. Martin, et al. - SNP 筛选
A SNaPshot assay for the rapid and simple detection of four common hotspot codon mutations in the PIK3CA gene.
Publication: BMC Research Notes 2009, 2:66
Authors: Hurst, D C,Zuiverloon, TCM, Hafner, C, Zwarthoff, EC et al. - 杂合性缺失
Multiplex SNaPshot Genotyping for Detecting Loss of Heterozygosity in the Mismatch-Repair Genes MLH1 and MSH2 in Microsatellite-Unstable Tumors.
Publication: Clin Chem 54(11): 1844-1854 (2008)
Authors: Bujalkova, M., K. Zavodna, et al. - 逆转录病毒抗性突变
Pre-screening HIV-1 reverse transcriptase resistance mutations in subtype B patients using a novel multiplex primer extension assay.
Publication: Curr HIV Res (2009) 7(4): 398-409.
Authors: Jakobsen, M. R., A. Aggerholm, et al. - 评估猪的肉用性能
SNaPshot minisequencing and a panel of candidate genes for complex routine testing of meat performance traits in pigs.
Publication: Anim Biotechnol.2007;18(2):109-15.
Authors: Civánová K, Knoll A. - 降解的 DNA
Developing multiplexed SNP assays with special reference to degraded DNA templates.
Publication: Nature Protocols 2006;1(3):1370-8
Authors: Sanchez, JJ.& Endicott, P. - 病原抗性
Single nucleotide polymorphism and haplotypes in the IL10 region associated with HCV clearance
Publication: Genes Immun.2005 Jun;6(4):347-57.
Authors: Oleksyk TK, Thio CL, Truelove AL, Goedert JJ, Donfield SM, Kirk GD, Thomas DL, O'Brien SJ, Smith MW. - 线粒体 SNP 筛选
Mitochondrial DNA mutations in patients with postlingual, nonsyndromic hearing impairment
Publication: European Journal of Human Genetics (2005) 13, 26−33.
Authors: Jacobs HT, Hutchin TP, Käppi T, Gillies G, et al. - 痒病易感性筛查
Primer extension assay for prion protein genotype determination in sheep.
Publication: Molecular and Cellular Probes (2004), 18(1), 33-37.
Authors: Vaccari G, Conte M, Morelli L, Di Guardo G, et al. - 癌变过程中的基因改变
Role of COX-2, thromboxane A2 synthase, and prostaglandin I2 synthase in papillary thyroid carcinoma growth.
Publication: Mod Pathol.2005 Feb;18(2):221-7.
Authors: Kajita S, Ruebel KH, Casey MB, Nakamura N et al. - 拟南芥标记物
Establishment of a high-efficiency SNP-based framework marker set for Arabidopsis.
Publication: Plant J. 2003 Oct;36(1):122-40.
Authors: Törjék O, Berger D, Meyer RC, Müssig C, Schmid KJ, Rosleff Sörensen T, Weisshaar B, Mitchell-Olds T, Altmann T. - 免疫中的 SNP 验证
Single nucleotide polymorphisms and haplotypes in the IL10 region associated with HCV clearance.
Authors: Oleksyk TK, Thio CL, Truelove AL, Goedert JJ, et al.
SNP 定量
- 突变体定量测定
Putative precursor cancer cells in human colorectal cancer tissue.
Publication: Int J Clin Exp Pathol (2009)2(2): 154-162
Authors: Goranova, T. E., M. Ohue, et al. - 等位基因特异性基因表达
Cis-acting variation in the expression of a high proportion of gene in human brain.
Publication: Hum Genet.2003 Jul;113(2):149-53.Epub 2003 May 1.
Authors: Bray NJ, Buckland PR, Owen MJ, O'Donovan MC. - 等位基因特异性基因表达
Allelic variation in human gene expression
Publication: Curr Opin Oncol.2004 Jan;16(1):39-43.
Authors: Yan H, Zhou W. - SNP 等位基因频率定量测定
Universal, robust, highly quantitative SNP allele frequency measurement in DNA pools.
Publication: Hum Genet.2002 May;110(5):471-8.Epub 2002 Mar 23.
Authors: Norton N, Williams NM, Williams HJ, Spurlock G et al.
BAC 指纹图谱分析
- 高通量细菌人工染色体指纹图谱分析
High-throughput fingerprinting of bacterial artificial chromosomes using
Publication: Genomics 82 (2003) 378–389
Authors: Ming-Cheng Luo, Carolyn Thomas, Frank M. You, Joseph Hsiao
甲基化
- 检测甲基化状态
Capillary electrophoretic analysis of methylation status in CpG-rich regions by single-base extension of primers modified with N6-methoxy-2,6-diaminopurine
Publication: Analytical Biochemistry Volume 380, Issue 1, 1 September 2008, Pages 13-20.
Authors: Boyd, V.L. et al. - 甲基化状态
Single nucleotide extension technology for quantitative site-specific evaluation of metC/C in GC-rich regions.
Publication: Nucleic Acids Res (2005) 33(10): e95.
Authors: Kaminsky, Z. A., A. Assadzadeh, et al.
软件下载
- 目前无软件可供下载。
用于 SNP 基因分型的重点产品：
Visistat reference component
仅供科研使用，不可用于诊断目的。