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Infinium Assay for Large-scale SNP Genotyping Applications
Published on: November 19, 2013
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基因定型SNP和Indels:一种方法来提高基于高分辨率融 (HRM) 分析的应用程序的范围和灵敏度
Kajan Muneeswaran1, Umayal Branavan2, Varuni A de Silva3
1Department of Chemistry, Faculty of Science, University of Colombo, Sri Lanka; Department of Psychiatry, Faculty of Medicine, University of Colombo, Sri Lanka.
概括
这项研究引入了不同化片 (CADMA) 试验的修改竞争放大,以改进高分辨率化 (HRM) 基因定型. 这种新方法提高了单核酸多态度 (SNP) 的准确性,并在各种qPCR平台上进行了研究.
科学领域:
- 分子生物学分子生物学
- 遗传学 遗传学 是一个
- 生物技术是生物技术.
背景情况:
- 高分辨率融 (HRM) 分析是一种闭管方法,用于检测单核酸多态 (SNP).
- 人力资源管理在灵敏度和准确性方面存在局限性,特别是在中性序列变化 (III类,IV类SNP和小INDEL) 方面,通常需要专门的,昂贵的设备.
- 由于近邻中性变化,现有的HRM分析面临敏感度降低和高错误率的挑战.
研究的目的:
- 设计一种基于不同化片 (CADMA) 的改性竞争性放大试验,以进行可靠的HRM基因定型.
- 创建可在大多数qPCR平台上检测到的基因特异性qPCR产品,克服专用HRM设备的局限性.
- 为应对基于HRM的基因定型中各种SNP类别和indels所带来的挑战.
主要方法:
- 从所有四个类别和indels (1bp和19bp) 选择的SNP用于测试设计.
- 利用修改后的CADMA原理,生成具有不同化温度 (Tm) 的等位基特异性片.
- 在CFX96平台上执行qPCR和曲线获取,使用Precision Melt软件进行分析,并将Tm预测与在线工具进行比较.
主要成果:
- 通过修改的CADMA-HRM-qPCR试验,成功地基因型化了各种SNP和INDEL.
- 使用对照样本和验证的Tm预测识别了基因型集群.
- 观察到使用布莱克和德尔科特 (1998) 方法计算的Tm值,调整为盐度,与实验Tm值非常相匹配.
结论:
- 开发的修改后的CADMA方法显著降低了基于HRM的基因造型失败率.
- 这种技术适用于各种qPCR平台上的任何SNP或indel.
- 成功实施需要了解融仪器,目标特征 (SNP类,INDEL长度,GC含量),并使用控制.
相关概念视频
02:26Comparing Copy Number Variations and SNPs
Sequencing of the human genome has opened up several best-kept secrets of the genome. Scientists have identified thousands of genome variations that exist within a population. These variations can be a single nucleotide or a larger chromosomal variation.
Copy number variations or CNVs are the structural variations that cover more than 1kb of DNA sequence. The single nucleotide polymorphism (SNP), on the other hand, is a single nucleotide change or a point mutation that is found in more than 1%...
Copy number variations or CNVs are the structural variations that cover more than 1kb of DNA sequence. The single nucleotide polymorphism (SNP), on the other hand, is a single nucleotide change or a point mutation that is found in more than 1%...
01:05Single Nucleotide Polymorphisms-SNPs
A single nucleotide polymorphism or SNP is a single nucleotide variation at a specific genomic position in a large population. It is the most prevalent type of sequence variation found in the human genome. Point mutations that occur in more than 1% of the population qualify as SNPs. These are present once every 1000 nucleotides on an average in the human genome. Replacement of a purine with another purine (A/G) or a pyrimidine with another pyrimidine (C/T) is known as a transition. In contrast,...
01:11Genome-wide Association Studies-GWAS
Genome-wide association studies or GWAS are used to identify whether common SNPs are associated with certain diseases. Suppose specific SNPs are more frequently observed in individuals with a particular disease than those without the disease. In that case, those SNPs are said to be associated with the disease. Chi-square analysis is performed to check the probability of the allele likely to be associated with the disease.
GWAS does not require the identification of the target gene involved in...
GWAS does not require the identification of the target gene involved in...