Detection of self-incompatible oilseed rape plants (Brassica napus L.) based on molecular markers for identification of the class I S haplotype

Abstract

The selection of desirable genotypes with recessive characteristics, such as self-incompatible plants, is often difficult or even impossible and represents a crucial barrier in accelerating the breeding process. Molecular approaches and selection based on molecular markers can allow breeders to overcome this limitation. The use of self-incompatibility is an alternative in hybrid breeding of oilseed rape. Unfortunately, stable self-incompatibility is recessive and phenotype-based selection is very difficult and time-consuming. The development of reliable molecular markers for detecting desirable plants with functional self-incompatible genes is of great importance for breeders and allows selection at early stages of plant growth. Because most of these reliable molecular markers are based on discrimination of class I S-locus genes that are present in self-compatible plants, there is a need to use an internal control in order to detect possible PCR inhibition that gives false results during genotyping. In this study, 269 double haploid F2 oilseed rape plants obtained by microspore embryogenesis were used to verify the applicability of an improved PCR assay based on the detection of the class I SLG gene along with an internal control. Comparative analysis of the PCR genotyping results vs. S phenotype analysis confirmed the applicability of this molecular approach in hybrid breeding programs. This approach allows accurate detection of self-incompatible plants via a different amplification profile.

Genetic insights into the modification of the pre-fertilization mechanisms during plant domestication

“…SI systems prevent self-fertilization, which forces outcrossing and increases genetic diversity, which is useful for the breeding of hybrid varieties of economically important plant families. Accordingly, breeding programs towards introducing functional SI have been activated in many crops (Kaothien-Nakayama et al, 2010;Havlícková et al, 2014;Cheng et al, 2018;Xiao et al, 2019). In order to control mating, many advances have been made toward understanding the SI mechanisms.…”

Progress on deciphering the molecular aspects of cell-to-cell communication in Brassica self-incompatibility response

“…In this regard, the use of molecular marker technology for the S-haplotype identification is advantageous that it could be done on young plants and no need to have the S-allele plants in flowering phase (Brace et al 1994). Hence, the development and identification of molecular markers linked with self-incompatible genes are of utmost value for geneticist and plant breeders to discriminate between class-I and class-II S haplotypes (Havlickova et al 2014). Different kinds of molecular markers have been reported for the identification of S haplotypes in Brassica species by different groups of researchers (Table 1), although all the S haplotypes have not been identified yet.…”

Early in vitro selection of winter oilseed rape (Brassica napus L.) plants with the fertility restorer gene for CMS Shaan 2A via non-destructive molecular analysis of microspore-derived embryos

“…Three F 1 hybrids from crosses of Rf lines and three cultivars of oilseed rape were used as microspore donor plants for the in vitro androgenesis process in the culture of isolated microspores. Microspore culture technique, cotyledon sampling and related experiments were carried out according to Klíma et al (2004Klíma et al ( , 2008 and Havlíčková et al (2014) with modified concentration of a doubling agent (trifluralin, 5 μmol/l). Once embryos with well-developed cotyledons reached a length of 4-5 mm (Figure 1A), approx.…”

“…All plants except haploids were individually and tightly covered with sacks made of dense fabric, grown to seed maturity in isolation cages and harvested. Genomic DNA was extracted from dissected cotyledons using a modified CTAB method (Havlíčková et al 2014). The concentration and quality of DNA were determined with a BioSpec-nano spectrophotometer (Shimadzu, Japan).…”

“…Surprisingly, DNA isolation even from smaller embryos (2.0-3.0 mm) provided quite a sufficient amount of material for DNA extraction. However, it may be difficult to obtain such a small sample always in a non-destructive way, without the risk of damage to the embryo tip or loss of the sample during routine manipulation (Havlíčková et al 2014(Havlíčková et al , 2015. The length of the amplified fragment was approx.…”