GENETIC MARKERS ASSOCIATED WITH STRIGA GESNERIOIDES RESISTANCE AND SEED SIZES IN COWPEA [VIGNA UNGUICULATA (L.) WALP.] INBRED LINES

ABSTRACT
Production of cowpea is severely hampered by different races of the parasitic weed, Striga gesnerioides. Cultivation of Striga-resistant cowpea is the most reliable protocol to effectively combat the parasite. The current study reported the identification of genetic markers associated with multi-race-Striga resistance and seed size across the genome of cowpea RIL populations. The landrace GH3684 from Ghana was tested for resistance against all known races of Striga gesnerioides in West Africa. Pot experiments were used to test for resistance in each line against Striga populations collected from Northern Ghana (GH) and Nigeria (SG3). Seed size was measured in field trials. SSR and SNP markers were identified and used for phylogenetic analysis and genetic mapping. A genetic linkage map was constructed with QTL IciMapping. Segregation of SSR-1 marker with known association with the Striga resistance gene rsg3 was 100 % consistent with the cowpea phenotypes in SG3. This study indicated that the SG3 resistance gene is located at 12.60 cM away from that of the GH race. On the whole, 70 % of the inbred lines of cowpea were resistant to Striga in Ghana and the lines designated UCC-11, UCC-24, UCC-32, UCC-122, UCC-221, UCC-241, UCC-328 (best RIL candidates for release) and GH3684 were immune to all 7 known races of Striga in West Africa. SARC-LO2 had resistance to four races of Striga (SG2, SG4z, SG5 and SG6). The low genetic diversity and polymorphism information content suggest close genetic relatedness within the RIL population. One and five SNP marker(s) were found to be associated with cowpea seed size and Striga resistance respectively.


CHAPTER ONE
INTRODUCTION
This chapter is about the general introduction of the current work; background to the study, statement of the problem, justification, objectives and the associated hypothesis.

Cowpea (Vigna unguiculata (L). Walp) is a dicotyledonous leguminous food crop. It is also commonly referred to as crowder pea or lubia. Cowpea is known to have originated from West Africa close to six thousand years ago and widely cultivated in different parts of the world including Latin America, Southeast Asia and in the southern United States of America (Girija

Dhanavel, 2009). Cowpea is usually cultivated by small scale farmers and sometimes intercropped with maize or sorghum. Diversity of cowpeas exist in West Africa particularly, Nigeria, Southern Niger, Ghana, northern part of Togo, part of Burkina Faso, northern Benin and the North-Western part of Cameroon (Ng & Marechal, 1985). Cowpea is an important food crop for both man and animals (Davis et al., 1991). It improves soil fertility and serves as income for farmers and traders. The fibers are used for making fishing lines and have also been considered as a source of pulp to make good quality papers (Summerfield, Huxley & Steel 1985).

Cowpea production is suitable for subsistence farming systems in which low inputs are involved due to its ability to thrive on relatively poor soil (Pasquet, 2000; Pronaf, 2003). It has high level of adaptation due to its inherent ability to withstand drought, tolerate shade, and fix atmospheric nitrogen (Singh, 1997). Despite the huge potential of cowpea to ensuring food security and good soil nutrient turn over, several factors are known to affect its production. The low productivity of cowpea is particularly due to intense biotic pressure by insects and other pest.

Cowpea is attacked by a host of pathogens, most prominent is the parasitic flowering plant Striga gesnerioides, whose parasitism causes severe chlorosis, wilting and stunting of susceptible hosts, leading to yield losses estimated to be around millions of tonnes annually (Aggarwal & Ouedraogo, 1989; Muleba et al., 1997; Singh & Emechebe, 1997). Studies conducted in West Africa by Lane et al. (1996) revealed that there are five different races of the Striga gesnerioides designated as SG1 through SG5. In addition, Botanga and Timko (2006) identified SG4z and SG6 bringing the known races to seven. Various control measures, including cultural practices, chemical control, biological control and host plant resistance have been suggested (Dube & Olivier, 2001; Boukar, Kong, Singh, Murdock & Ohm 2004). No single method, however, seems to be fully adequate in the control of this parasite. One practice, host plant resistance appears to effectively and economically control the parasite in that, it is affordable to resource-poor farmers (Omoigui et al., 2007). The only challenge with this control method is that different cowpea varieties react differently to different races of the parasitic plant at different locations in West Africa (Lane et al., 1993; Singh, 2004). This fact was established when 2 cultivars (58-57 and Suvita-2) were found to be completely resistant to S. gesnerioides in Burkina Faso (IITA 1982, 1975) but proved to be heavily susceptible to Striga in Niger and Nigeria when regional trials were conducted on these cultivars, suggesting strain variation in S. gesnerioides (Aggarwal, 1985). Well-adapted high-yielding cultivars resistant to all races of S. gesnerioides are under development but not widely available (Singh, 2000).

One of the important desirable traits of cowpea in West Africa is large seed size (Drabo, Redden, Smithson, & Aggarwal, 1984; Langyintuo et al., 2003; Tchiagam et al., 2011; Egbadzor et al., 2014). However, much breeding objectives have not been directly focused on seed size compared with such traits as biotic and abiotic stress tolerance (Orawu et al., 2013). Highly specific seed market classes for cowpea and other grain legumes exist because grain is most commonly cooked and consumed whole. Size, shape, colour, and texture are critical features of these market classes and breeders target development of cultivars for market acceptance. Resistance to biotic and abiotic stresses that are absent from elite breeding material are often introgressed through crosses to landraces or wild relatives. When crosses are made between parents with different grain quality characteristics, recovery of progeny with acceptable or enhanced grain quality is problematic. Several cycles of backcrossing help recover elite characteristics including seed size. However, this process can be cumbersome and inefficient due to possible linkage drag and the polygenic nature of the trait. Thus genetic markers for grain quality traits can help in pyramiding genes needed for specific market classes. Allelic variation dictating the inheritance of seed size can be tagged and used to assist the selection of large seeded lines.

The aim of cowpea breeding and genetic improvement programmes around the world is to put together desirable agronomic traits with resistances to the major diseases, insect pests or parasites that afflict cowpea in agro-ecologically adapted cultivars (Timko et al., 2007; Timko & Singh, 2008).

Approximately, ten years is required to breed a superior improved line using traditional selection and hybridization strategies. The overall efficiency and effectiveness of cowpea improvement programmes can be facilitated by the knowledge of the genetic diversity available within local and regional germplasm collections (Hall, Cisse, Thiaw, Elaward, Ehlers & Ismail, 2003; Hegde & Mishra, 2009). At present, analysis of Simple Sequence Repeat (SSRs) has proven to be useful since these sequences, besides being abundant and distributed throughout eukaryotic genomes, are highly polymorphic, inherited co-dominantly and reproducible, with simple screening requirements (Dib et al., 1996). Until recently, SSRs have been considered as the marker system of choice for the majority of applications. However, recent advances in sequencing and genotyping technologies now permit generation of large sets of single nucleotide polymorphism (SNP) markers from relatively under studied crop species such as faba bean at an acceptable level of cost. As a consequence, SNPs have become more widely used due to high abundance and capacity to be multiplex-formatted for high-throughput genotyping. In addition, SNP discovery from transcribed regions of the genome provides the basis to establish a direct link between sequence polymorphism and putative functional variation.

The cowpea breeding programme in the Department of Molecular Biology and Biotechnology, University of Cape Coast has the aim to introgress Striga-resistance quantitative trait locus (QTL) into local susceptible lines as well as improve their seed sizes. This has led to the development of new cowpea recombinant inbred lines, which warrant adequate genetic analysis to elucidate potential genotypes for optimum utilization of the crop. Besides, there is a need to validate multi-Striga race resistance status and seed size of the advanced cowpea recombinant inbred lines using simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers for subsequent selection and release of novel varieties.

Statement of the Problem
The most important challenge to cowpea cultivation in the major production regions of Northern Ghana and sub-Saharan Africa as a whole, is infestation of variable races of Striga gesnerioides in drought prone areas under poor soil conditions. The variable races of Striga warrant multi-resistance gene in cowpea to widely control the parasite across the value chain and trade routes in sub-Saharan Africa. The stress imposed on cowpea by Striga causes yield loss of between 80 % - 100 % (Asare et al., 2010). Besides, consumer preference for specific seed sizes has also become an important selective trait in breeding.

Obviously, the lack of improved Striga resistant cowpeas for farmers to cultivate, most especially, those at Ghana‟s hub of production (Upper East, Upper West and the Northern regions) largely contributes to the continual importation of cowpea from neighboring countries including Togo, Benin, Nigeria and Burkina Faso. The need for cultivation of improved Striga-resistant cowpeas in the affected regions in Ghana to meet consumer preference is critical towards sustainable production and food security. Though advanced recombinant inbred lines (RILs) of cowpea have been developed in the Department of Molecular Biology and Biotechnology, University of Cape Coast, it has not been fully explored for improved multi-Striga-resistance trait to mitigate the infestation of the parasitic weed. Besides, only few SSR markers exist and no SNP markers have been identified for genetic analysis to select multi-Striga-race resistant genotypes of cowpeas with improved seed sizes among inbred lines of the crop.

Justification
Variations in RILs of cowpea may be associated with useful multi-race-Striga-resistant and seed size traits. The use of molecular markers together with phenotypic information could be a more reliable protocol to identify and select desirable traits in crop improvement. Indeed, morphological and agronomic data coupled with molecular markers may facilitate reliable selection of multi-race-Striga-resistant cowpea genotypes with improved seed size. SSR and SNP genotyping are promising platforms for providing plant breeders with the simplest, most useful and cost-effective services, which can be employed to analyze the advanced RILs of cowpea which have been developed in the Department of Molecular Biology and Biotechnology, University of Cape Coast. This will enhance identification of multi-race-Striga- resistance and seed size QTLs across the genome of the inbred lines of cowpea.

Main Objective
The main objective of this study was to assess multi-race-Striga-resistance and seed size traits as well as genetic variability across the genome of recombinant inbred lines of cowpea.

Specific Objectives
The specific objectives were to determine:

* SSR markers associated with Striga resistance across the genome of cowpea inbred lines and GH3684

* Multi-Striga-race resistance status of some cowpea inbred lines and GH3684

* SNP markers associated with Striga-resistance and seed size among cowpea inbred lines.

* Genetic relatedness among the cowpea inbred lines

Hypotheses
* Specific SSR markers are linked to Striga-resistance across the genome of cowpea inbred lines.

* Differential multi-Striga-race resistant traits exist among some cowpea genotypes

* Specific SNP markers are associated with Striga-resistance and seed size

* Genetic variations exist among cowpea inbred lines

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Item Type: Ghanaian Postgraduate Material  |  Attribute: 145 pages  |  Chapters: 1-5
Format: MS Word  |  Price: GH50  |  Delivery: Within 30Mins.
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