Indian Institute of Spices Research, Kozhikode, Kerala, India.
Division of Crop Improvement and Biotechnology, ICAR-Indian Institute of Spices Research, Kozhikode, Kerala, India.
Methods Mol Biol. 2021;2222:219-247. doi: 10.1007/978-1-0716-0997-2_13.
Understanding biology and genetics at molecular level has become very important for dissection and manipulation of genome architecture for addressing evolutionary and taxonomic questions. Knowledge of genetic variation and genetic relationship among genotypes is an important consideration for classification, utilization of germplasm resources, and breeding. Molecular markers have contributed significantly in this respect and have been widely used in plant science in a number of ways, including genetic fingerprinting, diagnostics, identification of duplicates and selection of core collections, determination of genetic distances, genome analysis, development of molecular maps, and identification of markers associated with desirable breeding traits. The application of molecular markers largely depends on the type of markers employed, distribution of markers in the genome, type of loci they amplify, level of polymorphism, and reproducibility of products. Among many DNA markers available, random amplified polymorphic DNA (RAPD) is the simplest, is cost-effective, and can be performed in a moderate laboratory for most of its applications. In addition, RAPDs can touch much of the genome and has the advantage that no prior knowledge of the genome under research is necessary. The recent improvements in the RAPD technique like arbitrarily primed polymerase chain reaction (AP-PCR), sequence characterized amplified region (SCAR), DNA amplification fingerprinting (DAF), sequence-related amplified polymorphism (SRAP), cleaved amplified polymorphic sequences (CAPS), random amplified microsatellite polymorphism (RAMPO), and random amplified hybridization microsatellites (RAHM) can complement the shortcomings of RAPDs and have enhanced the utility of this simple technique for specific applications. Simple protocols for these techniques are presented along with the applications of RAPD in genetic diversity analysis, mapping, varietal identification, genetic fidelity testing, etc.
在分子水平上了解生物学和遗传学对于剖析和操纵基因组结构以解决进化和分类问题变得非常重要。了解基因型之间的遗传变异和遗传关系是分类、利用种质资源和进行育种的重要考虑因素。分子标记在这方面做出了重要贡献,并以多种方式广泛应用于植物科学,包括遗传指纹分析、诊断、重复基因的鉴定和核心群体的选择、遗传距离的确定、基因组分析、分子图谱的开发以及与理想育种性状相关的标记的鉴定。分子标记的应用在很大程度上取决于所使用的标记类型、标记在基因组中的分布、它们扩增的基因座类型、多态性水平以及产物的可重复性。在许多可用的 DNA 标记中,随机扩增多态性 DNA (RAPD) 最简单、最具成本效益,并且在大多数应用中可以在中等实验室中进行。此外,RAPD 可以触及大部分基因组,并且具有不需要对研究中的基因组有先验知识的优点。RAPD 技术的最新改进,如任意引物聚合酶链反应 (AP-PCR)、序列特征扩增区 (SCAR)、DNA 扩增指纹图谱 (DAF)、序列相关扩增多态性 (SRAP)、切割扩增多态性序列 (CAPS)、随机扩增多态性微卫星标记 (RAMPO) 和随机扩增多态性杂交微卫星 (RAHM),可以弥补 RAPD 的缺点,并增强了该简单技术在特定应用中的实用性。本文介绍了这些技术的简单方案,以及 RAPD 在遗传多样性分析、图谱绘制、品种鉴定、遗传保真度测试等方面的应用。