Genetics and analysis of quantitative traits

Single nucleotide polymorphism (SNP) data can be obtained using one of the numerous uniplex or multiplex SNP genotyping platforms that combine a variety of chemistries, detection methods, and reaction formats. Kompetitive Allele Specific PCR (KASP) is one of the uniplex SNP genotyping platforms, and has evolved to be a global benchmark technology. However, there are no publications relating either to the technology itself or to its application in crop improvement programs. In this review, we provide an overview of the different aspects of the KASP genotyping platform, discuss its application in crop improvement, and compare it with the chip-based Illumina GoldenGate platform. The International Maize and Wheat Improvement Center routinely uses KASP, generating in excess of a million data points annually for crop improvement purposes. We found that (1) 81 % of the SNPs used in a custom-designed GoldenGate assay were transferable to KASP; (2) using KASP, negative controls (no template) consistently clustered together and rarely produced signals exceeding the threshold values for allele calling, in contrast to the situation observed using GoldenGate assays; (3) KASP’s average genotyping error in positive control DNA samples was 0.7–1.6 %, which is lower than that observed using GoldenGate (2.0–2.4 %); (4) KASP genotyping costs for marker-assisted recurrent selection were 7.9–46.1 % cheaper than those of the BeadXpress and GoldenGate platforms; and (5) KASP offers cost-effective and scalable flexibility in applications that require small to moderate numbers of markers, such as quality control analysis, quantitative trait loci (QTL) mapping in bi-parental populations, marker-assisted recurrent selection, marker-assisted backcrossing, and QTL fine mapping.

Single nucleotide polymorphism genotyping using Kompetitive Allele Specific PCR (KASP): overview of the technology and its application in crop improvement

Random amplified polymorphic DNA (RAPD) markers have been used for numerous applications in plant molecular genetics research despite having disadvantages of poor reproducibility and not generally being associated with gene regions. A novel method for generating plant DNA markers was developed based on the short conserved region flanking the ATG start codon in plant genes. This method uses single 18-mer primers in single primer polymerase chain reaction (PCR) and an annealing temperature of 50°C. PCR amplicons are resolved using standard agarose gel electrophoresis. This method was validated in rice using a genetically diverse set of genotypes and a backcross population. Reproducibility was evaluated by using duplicate samples and conducting PCR on different days. Start codon targeted (SCoT) markers were generally reproducible but exceptions indicated that primer length and annealing temperature are not the sole factors determining reproducibility. SCoT marker PCR amplification profiles indicated dominant marker like RAPD markers. We propose that this method could be used in conjunction with these markers for applications such as genetic analysis, bulked segregant analysis, and quantitative trait loci mapping, especially in laboratories with a preference for agarose gel electrophoresis.

Start Codon Targeted (SCoT) Polymorphism: A Simple, Novel DNA Marker Technique for Generating Gene-Targeted Markers in Plants

The importance of plant genetic diversity (PGD) is now being recognized as a specific area since exploding population with urbanization and decreasing cultivable lands are the critical factors contributing to food insecurity in developing world. Agricultural scientists realized that PGD can be captured and stored in the form of plant genetic resources (PGR) such as gene bank, DNA library, and so forth, in the biorepository which preserve genetic material for long period. However, conserved PGR must be utilized for crop improvement in order to meet future global challenges in relation to food and nutritional security. This paper comprehensively reviews four important areas; (i) the significance of plant genetic diversity (PGD) and PGR especially on agriculturally important crops (mostly field crops); (ii) risk associated with narrowing the genetic base of current commercial cultivars and climate change; (iii) analysis of existing PGD analytical methods in pregenomic and genomic era; and (iv) modern tools available for PGD analysis in postgenomic era. This discussion benefits the plant scientist community in order to use the new methods and technology for better and rapid assessment, for utilization of germplasm from gene banks to their applied breeding programs. With the advent of new biotechnological techniques, this process of genetic manipulation is now being accelerated and carried out with more precision (neglecting environmental effects) and fast-track manner than the classical breeding techniques. It is also to note that gene banks look into several issues in order to improve levels of germplasm distribution and its utilization, duplication of plant identity, and access to database, for prebreeding activities. Since plant breeding research and cultivar development are integral components of improving food production, therefore, availability of and access to diverse genetic sources will ensure that the global food production network becomes more sustainable. The pros and cons of the basic and advanced statistical tools available for measuring genetic diversity are briefly discussed and their source links (mostly) were provided to get easy access; thus, it improves the understanding of tools and its practical applicability to the researchers.

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Importance of Genetic Diversity Assessment in Crop Plants and Its Recent Advances: An Overview of Its Analytical Perspectives

Marker-assisted selection (MAS) refers to the use of molecular markers to assist phenotypic selections in crop improvement. Several types of molecular markers, such as single nucleotide polymorphism (SNP), have been identified and effectively used in plant breeding. The application of next-generation sequencing (NGS) technologies has led to remarkable advances in whole genome sequencing, which provides ultra-throughput sequences to revolutionize plant genotyping and breeding. To further broaden NGS usages to large crop genomes such as maize and wheat, genotyping-by-sequencing (GBS) has been developed and applied in sequencing multiplexed samples that combine molecular marker discovery and genotyping. GBS is a novel application of NGS protocols for discovering and genotyping SNPs in crop genomes and populations. The GBS approach includes the digestion of genomic DNA with restriction enzymes followed by the ligation of barcode adapter, PCR amplification and sequencing of the amplified DNA pool on a single lane of flow cells. Bioinformatic pipelines are needed to analyze and interpret GBS datasets. As an ultimate MAS tool and a cost-effective technique, GBS has been successfully used in implementing genome-wide association study (GWAS), genomic diversity study, genetic linkage analysis, molecular marker discovery and genomic selection under a large scale of plant breeding programs.

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Genotyping-by-sequencing (GBS), an ultimate marker-assisted selection (MAS) tool to accelerate plant breeding.

The development and use of molecular markers for the detection and exploitation of DNA polymorphism is one of the most significant developments in the field of molecular genetics. The presence of various types of molecular markers, and differences in their principles, methodologies, and applications require careful consideration in choosing one or more of such methods. No molecular markers are available yet that fulfill all requirements needed by researchers. According to the kind of study to be undertaken, one can choose among the variety of molecular techniques, each of which combines at least some desirable properties. This article provides detail review for 11 different molecular marker methods: restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), inter-simple sequence repeats (ISSRs), sequence characterized regions (SCARs), sequence tag sites (STSs), cleaved amplified polymorphic sequences (CAPS), microsatellites or simple sequence repeats (SSRs), expressed sequence tags (ESTs), single nucleotide polymorphisms (SNPs), and diversity arrays technology (DArT).

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An overview of molecular marker methods for plants

Genetic mapping (also known as linkage mapping or meiotic mapping) refers to the determination of the relative position and distances between markers along chromosomes. Genetic map distances between two markers are defined as the mean number of recombination events, involving a given chromatid, in that region per meiosis. Genetic map construction requires that the researcher develop appropriate mapping population, decide the sample size and type of molecular marker(s) for genotyping, genotype the mapping population with sufficient number of markers, and perform linkage analyses using statistical programs. The construction of detailed genetic maps with high levels of genome coverage is a first step for localizing genes or quantitative trait loci (QTL) that are associated with economically important traits, marker assisted selection, comparative mapping between different species, a framework for anchoring physical maps, and the basis for map-based cloning of genes. Highly reproducible, high throughput, codominant, and transferable molecular markers, especially developed from expressed regions, are sought to increase the utility of genetic maps. This article reviews the principles, requirements, and future prospects of genetic mapping in plants.

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Principles, requirements and prospects of genetic mapping in plants

Marker assisted backcrossing (MAB) is one of the most anticipated and frequently cited benefits of molecular markers as indirect selection tools in breeding programs. However, routine implementations of MAB in ongoing plant breeding programs are still scarce. Currently MAB of single gene is perhaps the most powerful approach that uses DNA markers effectively. Improvement of quantitative traits loci (QTLs) through MAB resulted to variable results ranging from limited success and/or even a failure to a few highly successful stories. A major constraint to the implementation of MAB in pragmatic breeding programs has been the high relative cost compared to conventional phenotypic selection. It is a popular misconception that a ‘DNA fingerprint’ is always to be preferred. To be useful to plant breeders, gains made from MAB must be more cost-effective than gains through traditional breeding or MAB must generate significant time savings, which justifies the additional cost involved. Currently, most national agricultural research systems (NARS) in Africa have either no or very limited facilities, skilled manpower, and financing for integrating molecular markers as part of their breeding programs. Therefore, conventional breeding methods remain the main option for NARS for many years to come, but targeted use of MAB may become a supplement if well-validated markers are developed or available through collaboration with the international agricultural research centers. This paper provides detail review of the current literature on MAB, including requirements and selected experimental results.

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Progress and prospects of marker assisted backcrossing as a tool in crop breeding programs

NERICA rices are interspecific inbred progeny derived from crosses between Oryza sativa × O. glaberrima. In this study, we evaluated 70 BC2 interspecific lines, developed by crossing a tropical japonica variety (WAB 56-104) as the recurrent parent to an O. glaberrima variety (CG 14) as the donor parent, followed by the use of anther culture to derive doubled haploids (DH) (26 lines) or eight generations of inbreeding to fix the lines (44 lines). Seven of these BC2 derived inbred lines have been released as NERICA 1-NERICA 7. This study examined the relative contribution of each parent and the extent of genetic differences among these 70 sister lines using 130 well-distributed microsatellite markers which cover 1725 cM of the rice genome. The average proportion of O. sativa recurrent parent genome was 87.4% (1,508 cM), while the observed average proportion of O. glaberrima donor genome was 6.3% (108 cM). Non-parental alleles were detected in 83% of the lines and contributed an average of 38 cM per line (~2.2% of genomic DNA). Lines that had undergone eight generations of inbreeding in the field contained significantly more non-parental alleles (av. 2.7%) compared to the DH lines (av. 1.3%) that were developed from BC2 anthers. Using both cluster and principal component analyses, two major groups were detected in these materials. The NERICA varieties (NERICA 1 to 7) clustered in one group while the remaining 63 lines clustered in another group, suggesting that the second group may offer significant opportunities for further selection and variety development.

https://www.ajol.info/index.php/ajb/article/view/57884/46254

Molecular profiling of an interspecific rice population derived from a cross between WAB 56-104 ( Oryza sativa ) and CG 14 ( Oryza glaberrima )

The Africa Rice Center (WARDA) has developed several interspecific rice varieties by crossing the high yielding Asian rice (Oryza sativa subsp. japonica) with the locally adapted African rice (Oryza glaberrima). Eighteen varieties were named with the prefix NERICA (New Rice for Africa) but their genetic difference and patterns of relationship is largely unknown. A total of 102 polymorphic microsatellite markers were used to genotype 18 NERICAs. A subset of seven NERICAs (NERICA 1 to 7) was further characterized for 10 agronomic traits. The microsatellites data revealed no genetic difference between NERICA 8 and 9. The absence of genetic distance and identical SSR haplotype distribution (banding pattern) observed between NERICAs 8 and 9 is highly likely to be due to lack molecular difference at the DNA level but the possibility for seed admixture remains to be explored. This study, however, revealed the presence of a wide range of genetic differences among all other NERICAs, with the highest being between NERICA 6 and 17. Cluster and principal component analyses of the SSR data revealed distinct separation of NERICA 1 to 7 from NERICA 8 to 18. The possible reasons for such separation and the implications for breeding programs are discussed

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M. N. Ndjiondjop

Papers

An overview of molecular marker methods for plants

Principles, requirements and prospects of genetic mapping in plants

Progress and prospects of marker assisted backcrossing as a tool in crop breeding programs

Molecular profiling of an interspecific rice population derived from a cross between WAB 56-104 ( Oryza sativa ) and CG 14 ( Oryza glaberrima )

Microsatellites and agronomic traits for assessing genetic relationships among 18 New Rice for Africa (NERICA) varieties