Construction of a genetic linkage map based on amplified fragment length polymorphism markers and development of sequence-tagged site markers for marker-assisted selection of the sporeless trait in the oyster mushroom (Pleurotus eryngii)

Abstract

A large number of spores from fruiting bodies can lead to allergic reactions and other problems during the cultivation of edible mushrooms, including Pleurotus eryngii (DC.) Quél. A cultivar harboring a sporulation-deficient (sporeless) mutation would be useful for preventing these problems, but traditional breeding requires extensive time and labor. In this study, using a sporeless P. eryngii strain, we constructed a genetic linkage map to introduce a molecular breeding program like marker-assisted selection. Based on the segregation of 294 amplified fragment length polymorphism markers, two mating type factors, and the sporeless trait, the linkage map consisted of 11 linkage groups with a total length of 837.2 centimorgans (cM). The gene region responsible for the sporeless trait was located in linkage group IX with 32 amplified fragment length polymorphism markers and the B mating type factor. We also identified eight markers closely linked (within 1.2 cM) to the sporeless locus using bulked-segregant analysis-based amplified fragment length polymorphism. One such amplified fragment length polymorphism marker was converted into two sequence-tagged site markers, SD488-I and SD488-II. Using 14 wild isolates, sequence-tagged site analysis indicated the potential usefulness of the combination of two sequence-tagged site markers in cross-breeding of the sporeless strain. It also suggested that a map constructed for P. eryngii has adequate accuracy for marker-assisted selection.

Fig 1

Genetic linkage maps of Pleurotus eryngii based on 294 AFLP markers, two mating type factors, and the sporeless trait. Underlines represent skewed markers (0.001 ≤ P < 0.05), and daggers represent loci with more than four markers.

Fig 1

Genetic linkage maps of Pleurotus eryngii based on 294 AFLP markers, two mating type factors, and the sporeless trait. Underlines represent skewed markers (0.001 ≤ P < 0.05), and daggers represent loci with more than four markers.

Fig 2

Locus of the sporeless mutation and closely linked markers. (Left) map constructed by AFLP analysis; (right) map constructed by BSA-AFLP analysis. Daggers represent loci with more than four markers. Bold indicates markers selected as candidate STS markers. The description of each AFLP marker is given in Materials and Methods.

Fig 3

(A) Validation of STS markers SD488-I and SD488-II; (B) validation of both STS markers by multiplex PCR. The numbers from 1 through 17 represent, respectively, PE2-1, PE2-5, PE2, NPE010, ATCC 36047, KX-EG-9, Germany, Nara no. 1, ATCC 90212, ATCC 90787, ATCC 90887, ATCC 90888, UNICORN, WC827, 514, Tottoki-1, and Tottoki-2. These strains are listed in Table 1. M, 100-bp molecular ladder.