Genetics of resistance to anthracnose and identification of AFLP and RAPD markers linked to the resistance gene in PI 320937 germplasm of lentil (Lens culinaris Medikus)

Abstract

Anthracnose, caused by Colletotrichum truncatum, is a major disease problem and production constraint of lentil in North America. The research was conducted to examine the resistance to anthracnose in PI 320937 lentil and to identify molecular markers linked to the resistance gene in a recombinant inbred line (RIL) population developed from a cross of Eston lentil, the susceptible parent, and PI 320937, the resistant parent. A total of 147 F(5:6) RILs were evaluated for resistance to anthracnose in the greenhouse using isolate 95B36 of C. truncatum. Bulked segregant analysis (BSA) strategy was employed and two contrasting DNA bulks were constructed based on greenhouse inoculation of F(5)-derived F(6) RILs. DNA from the parents and bulks were screened with 700 RAPD primers and seven AFLP primer combinations. Analysis of segregation data indicated that a major dominant gene was responsible for resistance to anthracnose while variations in the resistance level among RILs could be the influences of minor genes. We designate the major gene as LCt-2. MapMaker analysis produced two flanking RAPD markers OPEO6(1250) and UBC-704(700) linked to LCt-2 locus in repulsion (6.4 cM) and in coupling (10.5 cM), respectively. Also, three AFLP markers, EMCTTACA(350) and EMCTTAGG(375) in coupling, and EMCTAAAG(175) in repulsion, were linked to the LCt-2 locus. These markers could be used to tag the LCt-2 locus and facilitate marker-assisted selection for resistance to anthracnose in segregating populations of lentil in which PI 320937 was used as the source of resistance. Also, a broader application of the linked RAPD markers was also demonstrated in Indianhead lentil, widely used as a source of resistance to anthracnose in the breeding program at the Crop Development Centre, University of Saskatchewan. Further selection within the few F(5:6) lines should be effective in pyramiding one or several of the minor genes into the working germplasm of lentil, resulting in a more durable and higher level of resistance.

An intersubspecific genetic map of Lens

“…A low genetic variability and insufficient genetic information are the reasons why until recently genetic maps of this species consisted of a relatively small number of markers, mainly isozymes and restriction fragment length polymorphisms (RFLPs) that covered an also relatively small portion of the lentil genome (Havey and Muehlbauer 1989;Weeden et al 1992;Tahir et al 1993). The recent development of a large number of molecular markers, especially RAPDs (randomly amplified polymorphic DNA) and AFLPs (amplified fragment length polymorphisms), is now providing the basis for constructing saturated maps in this species (Eujayl et al 1997(Eujayl et al , 1998Rubeena et al 2003), which consequently enables for the localization of genes of agronomic interest and facilitates a faster improvement in breeding programs (Tullu et al 2003). Inter-simple sequence repeats (ISSRs) are a relatively new type of DNA marker that involves the direct use of microsatellite sequences as primers in PCR analyses (Gupta et al 1994) to obtain DNA markers.…”

Screening techniques and sources of resistance to foliar diseases caused by major necrotrophic fungi in grain legumes

“…Screening of lentil germplasm has been conducted under field conditions as well as greenhouse conditions Tar'an et al, 2003a,b;Tullu et al, 2003Tullu et al, , 2005. In both approaches, artificial inoculation was employed.…”

“…Plants are inoculated with single isolate suspensions of 10 5 conidia ml −1 and 1.5 ml per plant 3-4 weeks after planting (10-12-node stage or early flowering). To avoid cross-contamination in experiments where different isolates are used , or where humidity around the plants has to be increased (Tullu et al, 2003), polyethylene wrapping is used. Plants are incubated for 24 h at 100% humidity, and then maintained in the greenhouse with 12-h supplemental light.…”

“…Controlled conditions. For greenhouse experiments, accessions are planted in 9 or 10 cm pots together with a susceptible and resistant control plant at a density of six to eight seeds per pot, using three pots as replicates Tar'an et al, 2003a,b;Tullu et al, 2003Tullu et al, , 2005. Plants are inoculated with single isolate suspensions of 10 5 conidia ml −1 and 1.5 ml per plant 3-4 weeks after planting (10-12-node stage or early flowering).…”

Plant Responses to Elevated Temperatures: A Field Study on Phenological Sensitivity and Fitness Responses to Simulated Climate Warming

“…Earlier studies have used RFLP, AFLP, and RAPD markers to assess genetic diversity and phylogenetic analyses within and among Lens species (Havey and Muehlbauer, 1989;Aboelwafa et al, 1995;Sharma et al, 1995Sharma et al, , 1996Ahmad and McNeil, 1996;Ford et al, 1997) and gene mapping (Eujayl et al, 1998b;Tullu et al, 2003;Duran et al, 2004;Kahraman et al, 2004;Hamwieh et al, 2005). As a part of the CGIAR's Generation Challenge Program (GCP), ICARDA has identified a composite collection of lentil germplasm and characterized them by using SSR markers.…”