Anther-mediated haploid induction and genetic stability of Phytophthora-resistant backcross lines of Capsicum annuum L

Abstract

Bell pepper is an important vegetable crop from Solanaceae family, which is cultivated and consumed globally due to its delicious, pleasant and nutrient-rich nature. Most of the commercially cultivated bell pepper cultivars are however, highly susceptible to P. capsici. They are recalcitrance to in vitro regeneration, which make the crop improvement efforts difficult through plant tissue culture-based approaches such as double haploid production. In this study, in vitro callus regeneration protocol was established from anthers of two Phytophthora resistant-backcross lines derived from crossing of two highly susceptible bell pepper cultivars (California Wonder and Solan Bharpur) with a highly resistant chilli landrace, CM334.The cultures were initially established in full strength MS medium, supplemented with different concentration of auxin and cytokinin. Maximum callus induction was achieved in MS medium CI2 supplemented with IAA (2 mg/L) and BAP (0.3 mg/L) (41.59 %) at pH 5.8, without heat-shock and cold stress treatments. The light-green and white compact calli were obtained which proliferated and maintained in the half strength MS medium containing 0.1 mg/L Kinetin (pH 5.8). The presence of Phytophthora resistance loci and successful crossings were confirmed through PCR amplification with the closely linked (RGA-C) and pungency −specific (Pun2) molecular markers, and phenotypic evaluations against P. capsici. The haploid nature and genetic stability were revealed through squash staining with 2 % acetocarmin. This study standardized the conditions appropriate for double haploid culture and regeneration of whole plants to speed up the development of Phytophthora-resistant bell pepper varieties.