Rapid identification of 1-aminocyclopropane-1-carboxylate (ACC) synthase genotypes in cultivars of Japanese pear (Pyrus pyrifolia Nakai) using CAPS markers

Abstract

In Japanese pear (Pyrus pyrifolia Nakai), fruit storage potential is closely related to the amount of ethylene produced. We have developed a rapid and accurate method for analyzing genes involved in high ethylene production during fruit ripening in Japanese pear. This involves cleaved-amplified polymorphic sequences (CAPS) of two 1-aminocyclopropane-1-carboxylate (ACC) synthase genes (PPACS1 and PPACS2). Two CAPS markers (A for PPACS1 and B for PPACS2), associated with the amount of ethylene produced, were identified. Marker A was associated with high ethylene producers and marker B with moderate ethylene producers. The absence of these two markers enabled the identification of low ethylene producers. Using these markers, we have identified ethylene genotypes for 40 Japanese pear cultivars and two Chinese pear (P. bretschneideri) cultivars that are commercially important and used in breeding programs. Furthermore, we performed linkage analysis of these two genes in the F(2) population, which revealed that the recombination frequency between the two markers was 20.8 +/- 3.6%. This information is critical to the selection of parents and in breeding strategies to improve storage ability of Japanese pears.

Genotyping-by-sequencing of pear (Pyrus spp.) accessions unravels novel patterns of genetic diversity and selection footprints

“…GWA analysis in this study identified a new QTL for fruit firmness on LG16, and the next best SNP explaining about 5% of variation in fruit firmness was located on LG15, where two ethylene producing genes ( PpACS1 and PpACS2 ) were identified in an earlier study. 61 The most significant SNP, explaining 8% of variation in TA, was located on LG2, which is the same one as that of an earlier study using a bi-parental cross between European and Asian species. 62 The next best SNP associated with TA was located on LG7 (Supplementary Figure S3), which appears to be a new genomic region that has not been reported before.…”

Potential assessment of genome-wide association study and genomic selection in Japanese pear <i>Pyrus pyrifolia</i>

“…Itai et al (2003) found that PPACS2 is associated with moderate ethylene production, suggesting that the association of PPACS2 observed in the present study is related to the ethylene production ability of Japanese pear cultivars. Although Itai et al (2003) also found that another marker, PPACS1, was strongly associated with ethylene production, we found no significant association between PPACS1 and harvest time in the present analysis. This is probably because PPACS1 did not have causal alleles in the population of the 76 Japanese pear cultivars.…”

“…In Japanese pear, ripening and fruit storage potential are related to the amount of ethylene produced. Harvest time is related to the maximum ethylene production during fruit ripening (Itai et al 2003). ACC synthase (1-aminocyclopropane-1-carboxylate synthase) converts S-adenosyl methionine (SAM) to ACC.…”

“…The 76 varieties were genotyped for 162 DNA markers, including 155 simple sequence repeats (SSRs) from pear and apple, four RAPD-STS, two ACC synthase genes and one locus encoding S-RNase gene, which are listed in the Appendix (Guilford et al 1997, Ishimizu et al 1999, Itai et al 2003, Liebhard et al 2002, Sawamura et al 2004, Silfverberg-Dilworth et al 2006, Terakami et al 2006, Yamamoto et al 2002b). Then SSR-PCR amplification was performed in a 10 μl reaction solution containing 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM MgCl2, 0.001% gelatin, 0.2 mM each of dNTPs, 5 pmol of each forward primer labeled with fluorescent chemical (Fam/Vic/Ned) and unlabelled reverse primer, 5 ng of genomic DNA and 0.25 unit of Taq polymerase (Invitrogen Corp., USA).…”

“…Four RAPD-STS markers were analyzed according to the method described in Terakami et al (2006). Two ACC synthase genes, which were involved in the ethylene synthesis pathway, were genotyped using the method reported by Itai et al (2003). Linkage groups and roughly estimated map positions of DNA markers in this study were identified using three genetic linkage maps of a Japanese pear variety ‘Hosui’ and European pear varieties ‘Bartlett’ and ‘La France’ (Nishitani et al 2009, Terakami et al 2009, Yamamoto et al 2007).…”