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QTLs associated with resistance to the cassava mosaic disease

Y. Lokko1, Melaku Gedil2 and Alfred Dixon1

1International Institute of Tropical Agriculture (IITA), Oyo Road PMB 5320, Ibadan, Nigeria, www.iita.org,
emails y.lokko@cgiar.org, A.Dixon@cgiar.org .
2
Georgetown University Medical Centre, 3900 Reservoir Road NW, Washington, DC 20007-2197 USA
email: mag68@georgetown.edu

Abstract

The cassava mosaic disease (CMD) is an economically important disease of the food crop cassava (Manihot esculenta Crantz) in Africa. Quantitative trait loci (QTLs) associated with resistance to CMD were identified using 132 F1 progeny derived from a cross between the CMD resistant accession TMSI30572 and the susceptible landrace TME117. The population was evaluated in the field for two growing seasons in Nigeria. Mean disease severity scores (MDSS) were used for QTL analysis. Five highly significant (p<0.001) marker-associated QTL effects, explaining between 10.47 and 12.15% of the total phenotypic variation, were detected by regression in the old source of resistance. The Kruskal Wallis analysis further detected four highly significant marker-associated QTL effects. Significant marker trait associations were due to markers donated by both parents, which confirms the polygenic and recessive nature of this source of resistance.

Media summary

Quantitative trait loci (QTLs) associated with resistance to the cassava mosaic virus disease (CMD) from the main source of resistance to CMD in Africa were identified.

Keywords

Linkage map, Marker assisted selection, Simple sequence repeats

Introduction

The cassava mosaic virus disease (CMD), which is caused by one of four distinct cassava begomoviruses, is the most important disease of cassava in Africa (Geddes, 1990). Earlier breeding efforts in the 1920’s introgressed resistance from Manihot glaziovii Muller von Argau into cassava (Nichols, 1947). Clone 58308, which was selected from this process and its progenies, has been the main source of breeding was for resistance to CMD. This source of resistance is recessive and polygenic (Hahn, et al., 1980). Resistance to CMD has also recently been identified among the landraces. Genetic studies have shown that the new sources of resistance in the landraces are either polygenic and recessive, or exhibit single dominant inheritance (Lokko et al 1998; Akano et al., 2001). Further progress in understanding CMD resistance would be made by identifying the individual gene or genes affecting resistance. The detection of multiple genes for virus resistance using segregation analysis alone is not efficient because of the differences due to genotype by environment interaction (McMullen and Louie 1989). Molecular markers, which are not affected by environmental conditions and are insensitive to gene interactions, are suitable for such studies. Markers found to be associated with genes for resistance could also be developed for marker-assisted selection (MAS) to facilitate breeding.

The objective of this study was to identify QTLs associated with resistance to CMD in the old sources of resistance to the disease using simple sequence repeat markers (SSRs).

Materials and Methods

Plant material

Segregating F1 progenies from a cross between the breeder’ accession TMS I30572 and landrace TME117 (Isunikankiyan) were established from embryonic axes, multiplied and six copies of each genotype were transferred to a seedling nursery in Abuja, Nigeria, which is a low pressure site for CMD. TMS I3072, which exhibits resistance to CMD, is from the earlier breeding selection and is an off spring (open pollinated seed) of a cross between 58308 and the South American accession, Branca de Santa Caterina, while TME117 is from Oyo State, Nigeria and is susceptible to CMD.

Phenotypic screening

Cuttings were made from each genotype and planted at Onne Nigeria, a high humid forest agro ecology site and a high pressure site for CMD, and were evaluated during the 1998/1999 and 1999/2000 seasons. Each trial had three replicates with 10 plants of a genotype in each replicate. Individual plants of all genotypes were assessed at 1, 3, 6 and 9 months after planting (MAP) for their reaction to CMD under natural infection by whiteflies. CMD severity was assessed using the standard CMD scoring scale of 1 to 5 where 1= no visible symptom and 5= very severe symptoms and stunting of the entire plant. Data was subjected to GLM analysis and mean disease severity score (MDSS) of the individual genotypes were computed using least square means.

Genetic Linkage Mapping

Following DNA extraction from the parents and progenies using the standard Dellaporta method (Dellaporta, et al 1983), the parents were screened with 349 cassava SSR primers, then the polymorphic primers were used to genotype the population. Linkage analysis was performed on the data generated using MAPMAKER version 3.0 computer programme (Lander et al., 1987).

QTL analysis

QTL analysis was performed on the mean disease severity scores (MDSS) derived by least square means from all the marker data. Marker trait association for resistance to CMD was determined by linear regression using the SAS computer package (SAS, 1999). Marker association was further validated with the non-parametric mapping procedure (Kruskal Wallis rank-sum test) of the software package MapQTL versions 4.0 (Van Ooijen et. al. 2002).

Results

A total of 125 markers were subjected to linkage analysis at a likelihood of odds (LOD) score of 3.0 and recombination fraction of 0.18 as the threshold for declaring linkage. Sixty-two markers mapped in 19 linkage groups (LGs), which covered 816.6 cM of the genome (Fig 1). Single marker analysis by regression yielded two significant (P<0.001) marker associated effects on LG XI and an unmapped marker donated by the resistant parent. One significant (P<0.001) marker associated effect on LG XVII and one on LG XVIII were donated by the susceptible parent. These markers explained 12.15, 11.55%, 10.64%, 10.47% and 11.30% of the phenotypic variation respectively (Table 1). The Kruskal-Wallis test further revealed four significant (P<0.001) markers, including three detected by regression, and a marker donated by the susceptible parent, which was not detected by regression. A fifth significant (P<0.005) marker associated effect, donated by the resistant parent, was detected by the Kruskal-Wallis test. Generally, the CMD resistance associated markers donated by either the resistant or susceptible parent mapped on different LG. Significant marker trait associations due to markers donated by both the resistant and susceptible parents, implies that both parents contributed to resistance in their progeny. This is consistent with the results from quantitative genetic studies, which suggest that there are multiple genetic factors for resistance to CMD in Casava and the resisatnce to CMD from clone 58303 and its progeny is recessive (Hahn, et al., 1980).

Table 1. Association between markers and CMD resistance in mapping population TMS I30572 X TME117 based on linear regression and Kruskal Wallis rank-sum test (K)

Locus

Donating parent

LG

Map Position

R2

K

SSRY77

TME117

XVII

19.9

 

11.97*****

SSRY7

TME117

XVIII

0

11.30****

10.66****

SSRY6p1b

TMS I30572

XI

10.9

12.15****

8.20****

SSRY21p2

TME117

XVII

40.1

10.47****

9.30****

SSRY42p1a

TMS I30572

XI

0

11.55****

 

SSRY324

TMS I30572

UM

 

10.64****

 

SSRY21p1

TMS I30572

UM

   

7.31***

*** p< 0.01 probability level
**** p< 0.005 probability level
***** p< 0.001 probability level
LG Linkage group

Conclusion

The efficiency of MAS for CMD resistance using these markers associated with the QTLs identified in this study still needs to be investigated. Additional markers are required to obtain dense maps. The inclusion of additional markers based on sequenced data such as ESTs and RFLPs converted to PCR-based markers onto the present TMS I30572 X TME117 framework map would provide better coverage of the genome and increase their usefulness in gene tagging and MAS of resistance to CMD and other traits of agronomic importance.

References

Akano, A.O., A.G.O. Dixon, C. Mba, E. Barrera and M. Fregene, 2002. Genetic mapping of a dominant gene conferring resistance to the cassava mosaic disease (CMD). Theoretical and Applied Genetics 105:521–525.

Dellaporta SL Wood J, Hicks JR (1983) A plant DNA minipreparation: version II. Plant Molecular Biology Reporter 1:19-21

Geddes, A. M., 1990. The relative importance of crop pests in sub-Saharan Africa. Bulletin 36; Chatham Natural Resource Institute (NRI).

Hahn, S.K., E.R. Terry and K. Leuschner, 1980. Breeding cassava for resistance to cassava mosaic virus disease. Euphytica 29:673-683

Lander, E.S., P. Green, J. Abrahamson, A. Barlow, M.J. Daly, S.E. Lincoln, and L. Newberg, 1987. MAPMAKER: An interactive computer package for constructing primary genetic linkage maps of experimental and natural populations. Genomics 1:174-181.

Lokko, Y., A.G.O. Dixon and S.K. Offei, 1998. Combining Ability of resistance to the cassava mosaic virus disease. In M.O. Akorada and J.M. Ngeve (Eds). Root Crops in the 21st Century. Proceedings of the 7th Triennial conference of the International Society for Tuber and Root Crops, Africa Branch,. 11 –17 Oct. 1998 Cotonou, Benin, pg 438-442. IITA Ibadan, Nigeria.

McMullen, M.D. and R. Louie. 1989. Linkage of molecular markers to a gene controlling the symptom response in maize to maize dwarf mosaic virus. Molecular plant. Microbe Interaction 2(6):309-314..

Nichols, R.F.W., 1947. Breeding cassava for virus resistance. East African Agricultural Journal 15:154-160.

SAS Institute, 1999. SAS companion for Microsoft windows environment, version 6, 1st ed. SAS Institute, Cary, North Carolina, USA.

Fig 1 Genetic map of population A TMS I30572XTME117. Markers labeled in green are donated by susceptible male parent TME117

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