A rapid real‐time PCR assay for detecting columnar growth in apples (Malus × domestica)

Abstract

Columnar growth in apples can have considerable positive consequences for orchard maintenance, but the lack of columnar cultivars producing fruits with desired market quality prevents their broader utilization in orchards. New crossings focused on their breeding are therefore conducted throughout the world every year. Unfortunately, the reliable discrimination between columnar growth and normal phenotype is difficult until seedlings are 2-3 years of age. However, molecular marker-assisted selection (MAS) can be used in seedlings several weeks old providing immense help in breeding programmes focused on columnar apple production. Here, we describe a rapid and reliable method for the detection of both wild-type (WT) and columnar (Co) alleles originating from 'McIntosh Wijcik' in a single reaction using a real-time PCR allelic discrimination approach. Our assay was tested in 130 genotypes (80 normal and 50 columnar habit) from various crossings and their parents, and in all cases, genotype corresponded to phenotype. The assay is thus suitable for laboratories interested in high-throughput MAS for columnar tree habit breeding.

Molecular Mechanisms Regulating the Columnar Tree Architecture in Apple

“…Columnar apples have labor-saving properties for apple growers and fit well with sustainable high-density planting, resulting in higher yields per hectare. However, an important problem is that columnar apple cultivars with superior fruit quality have not yet been developed [48]. The columnar growth phenotype can also easily be combined with other desirable characteristics (e.g., disease resistance) by crossing because the columnar growth phenotype is controlled by the dominant Co gene [47].…”

“…It is difficult to differentiate between columnar and non-columnar apples phenotypically until the seedlings are 2-3 years old [48]. Therefore, MAS systems, which can be applied to several-week-old seedlings, facilitate the selection of columnar apples.…”

“…However, these DNA markers could not discriminate between heterozygous columnar apples (Co/co) and homozygous columnar apples (Co/Co) when a single PCR was performed. Okada et al [25] and Cmejlova et al [48] conducted PCRs using three primers designed from the insertion polymorphism and simultaneously identified homozygous columnar apples (Co/Co), heterozygous columnar apples (Co/co), and non-columnar apples (co/co) through a single PCR. These Co allele-specific PCR products are useful DNA markers in the MAS of columnar apples for segregating progenies even when 'McIntosh' or its offspring are used as a parent because it can distinguish the Co allele from the original wild-type co allele [25,26].…”