Correlation of stages of microsporogenesis with bud and anther morphology in pepper genotypes through DAPI staining with different levels of mordant in cytological fixative.

The haploid and doubled haploid plants serve as valuable tools for breeders due to their ability to expedite the mapping of genes of agronomic importance, as well as accelerate the breeding cycle for generation of novel hybrids and improved homogenous varieties. Successful anther/microspore culture largely depends on the use of microspores at appropriate developmental stages at the time of culture, which can be specific for each plant species and genotype. In the present study, we described the visible morphological characteristics of flower buds and anthers at different developmental stages to identify the optimal microspore stage within the anther/buds of two pepper hybrids, Indra and Lakshmi. This information enabled us to predict the suitable microspore stage for successful haploid production. To enhance the visualization of nuclei in the pepper microspores, different concentrations of FeCl3 were employed as a mordant to Carnoy's fixative I, followed by DAPI staining. A clear and distinct nucleus was observed using DAPI staining procedures in the pepper microspores when fixed in Carnoy's solution containing ferric chloride (40-90 µl) as mordant. The use of mordant thus facilitated the efficient cytological analysis of the pepper microspores. Present results indicate that, to achieve efficient haploid production, flower buds with an average length of 4.4 to 5.02 mm for the hybrid Indra and 5.15 to 5.40 mm for the hybrid Lakshmi should be utilized. Additionally, these buds should have a calyx covering approximately 80-90% of the total bud length. We observed that in such buds, microspores are in the late-uninucleate and early binucleate stage which has been reported to be the most conducive stage for androgenesis induction in pepper.

Integrating marker-assisted background analysis with foreground selection for pyramiding bacterial blight resistance genes into Basmati rice.

Bacterial leaf blight (BB), caused by the bacterium Xanthomonas oryzae pv. Oryzae (Xoo), is the major constraint amongst rice diseases in India. CSR-30 is a very popular high-yielding, salt-tolerant Basmati variety widely grown in Haryana, India, but highly susceptible to BB. In the present study, we have successfully introgressed three BB resistance genes (Xa21, xa13 and xa5) from BB-resistant donor variety IRBB-60 into the BB-susceptible Basmati variety CSR-30 through marker-assisted selection (MAS) exercised with stringent phenotypic selection without compromising the Basmati traits. Background analysis using 131 polymorphic SSR markers revealed that recurrent parent genome (RPG) recovery ranged up to 97.1% among 15 BC3F1 three-gene-pyramided genotypes. Based on agronomic evaluation, BB reaction, aroma, percentage recovery of RPG, and grain quality evaluation, four genotypes, viz., IC-R28, IC-R68, IC-R32, and IC-R42, were found promising and advanced to BC3F2 generation.

CRISPR/Cas9: a promising way to exploit genetic variation in plants.

Creation of variation in existing gene pool of crop plants is the foremost requirement in crop improvement programmes. Genome editing is a tool to produce knock out of target genes either by introduction of insertion or by deletion that disrupts the function of a specific gene. The CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) system is the most recent addition to the toolbox of sequence-specific nucleases that includes ZFNs and TALENs. The CRISPR/Cas9 system allows targeted cleavage of genomic DNA guided by a small noncoding RNA, resulting in gene modifications by both non-homologous end joining and homology-directed repair mechanisms. Here, we present an overview of mechanisms of CRISPR, its potential roles in creating variation in germplasm and applications of this novel interference pathway in crop improvement. The availability of the CRISPR/Cas9 system holds promise in facilitating both forward and reverse genetics and will enhance research in crops that lack genetic resources.