Drought stress at the reproductive stage in rice is one of the most important cause for yield reduction, affecting both productivity and quality. All Basmati rice varieties, including the popular cultivar ‘Pusa Basmati 1 (PB1)’ is highly sensitive to reproductive stage drought stress (RSDS). We report for the first time, improvement of a Basmati cultivar for RSDS tolerance, with the introgression of a major quantitative trait locus (QTL), ‘qDTY1.1’ into PB1. The QTL donor was sourced from an aus variety, Nagina 22 (N22). A QTL linked microsatellite (SSR) marker ‘RM 431’ was employed for foreground selection for qDTY1.1 in the marker assisted backcross breeding process. A set of 113 SSR markers polymorphic between N22 and PB1 were utilized for background selection to ensure higher genome recovery. After three backcrosses followed by five generations of selfing, eighteen near isogenic lines (NILs) were developed, through combinatory selection for agro-morphological, grain and cooking superiority traits. The NILs were evaluated for three consecutive Kharif seasons, 2017, 2018 and 2019 under well-watered and drought stress conditions. RSDS tolerance and yield stability indicated that NIL3, NIL5, NIL6, NIL7, NIL12, NIL15 and NIL17 were best in terms of overall agronomic and grain quality under RSDS. Additionally, NILs exhibited high yield potential under normal condition as well. The RSDS tolerant Basmati NILs with high resilience to water stress, is a valuable resource for sustaining Basmati rice production under water limiting production environments.

The generation of isogenic induced pluripotent stem cell (iPSC) lines using CRISPR-Cas9 technology is a technically challenging, time-consuming process with variable efficiency. Here we use fluorescence-activated cell sorting (FACS) to sort biallelic CRISPR-Cas9 edited single-cell iPS clones into high-throughput 96-well microtiter plates. We used high-content screening (HCS) technology and generated an in-house developed algorithm to select the correctly edited isogenic clones for continued expansion and validation. In our model we have gene-corrected the iPSCs of a Parkinson’s disease (PD) patient carrying the autosomal dominantly inherited heterozygous c.88G>C mutation in the SNCA gene, which leads to the pathogenic p.A30P form of the alpha-synuclein protein. Undertaking a PCR restriction-digest mediated clonal selection strategy prior to sequencing, we were able to post-sort validate each isogenic clone using a quadruple screening strategy. Subsequent transfection with mRNA encoding excision-only transposase allows for the generation of footprint-free isogenic iPSC lines. These monoclonal isogenic iPSC lines retain a normal molecular genotype, express pluripotency markers and have the ability to differentiate into the three germ layers. This combinatory approach of FACS, HCS and post-sorted restriction digestion facilitates the generation of isogenic cell lines for disease modelling to be scaled-up on an automated platform.