Precision in transfer and editing of target gene is of more significance in genetic manipulation of crop plants. The target gene GhPHYA1 is one of the potential major negative regulators of many genes contributing for cotton fiber quality, boll size, vegetative growth, flowering etc. Therefore, targeted mutagenesis of GhPHYA1 aimed at improvement of useful trait in cotton.

Agrobacterium-mediated genetic transformation of G. hirsutum Coker 312 with CICR- cry2Ab1Ac::chitinase gene constructs and regeneration through somatic embryogenesis found four putative transgenic plants positive for npt-II and chitinase gene with PCR analysis using gene specific primers and 45 putative transgenic plants were regenerated.

Characterization of WNT-like gene/s in plants and functional validation may leads to unveiling their potential role in somatic embryogenesis of cotton. Such an attempt of basic research not only aids for the generation of reproducible somatic embryogenesis protocol but also may create clues for the signalling cascades of WNT-like pathways to uncover other upstream or downstream players in plants.

Putative transgenic callus cultures derived from transformation of Wnt 3A gene cassette in G hirsutum cv. Suraj were confirmed through PCR analysis using combination of gene and vector backbone primers. Isolation, cloning and transformation of Wnt 3A like gene into cotton (Suraj) was attempted for its functional validation through over-expression.

Introgression and Evaluation of Tg2E13 event: Embryo culture mediated accelerated introgression of Tg2E13 event (crylAc gene) into three genotypes (Suraj, NH615 and CISH3178) was accomplished through backcross breeding under contained facility of ICAR-CICR, Nagpur. To confirm the protein expression, bioefficacy and agronomic performance of the Tg2E13 introgressed plants, a Confined Field Trial was proposed to conduct in the year 2021 -22.

Introgression of CH12 event (cry2Ax1): BC3F,’s of three crosses (Suraj, NH615 and CISH3178) were made, F1’s will be selfed to produce BC3F2. It is proposed to study homozygous plants in BC3F2 generation and shall be further evaluated.

Evaluation of Event UASD 78 (cry1Ac): The event UASD 78 has been characterized and the report was submitted to the Council.

Evaluation of Tma12 (Event 403) on whitefly incidence: Comparative evaluation of Event 403 (Tma-12), non-Event, HS-6 (susceptible genotype) and LPS-141 (resistant genotype) was carried out in 2020-21. Incidence of whitefly on Tma 12 and LPS141 (resistant check) was similar while the incidence on non-Tma 12 plants and HS6 (highly susceptible check) was comparable but significantly higher than in Tma12 and LPS141 genotypes.

The deployment of biotechnology tools for cotton crop improvement necessitates the discovery of novel genes associated with economical important traits and unveiling the possible role of engineered Cry toxin for enhanced toxicity against cotton bollworms. The cry1D nucleotide sequence coding for active toxin was custom synthesized, cloned in bacterial expression vector pET28, expressed in bacterial host E. coli Rosetta garni and BL21 strains.

Recombinant proteins expressed in E. coli BL21 cells were isolated and utilized for testing the efficacy of target Cry1D protein against P. gossypiella, H. armigera and S. frugiperda through diet incorporation method. The preliminary bioassay results were found promising against pink bollworm P. gossypiella.

Isolation of stress (drought and salt) responsive genes for functional validation: Isolated full length coding sequence of GaDARI (1.5kb) and GhNAC9 (861 bp) from G. arboreum Cv Roja and G. hirsutum respectively.

PHYA1 is one of the known major potential negative regulator of PHYA2, PHYB PHYC and PHYE genes and RNAi cotton lines of PHYA1 showed vigorous vegetative growth, flowering and early maturity, higher root mass, higher germination, more flowers and bolls, and increase in fibre staple length.

ghPHTAI Chimeric callus cultures for the presence of sgRNA::Cas9 using vector and sgRNA specific primers was established. This methodology helps to shortlist the putative callus cultures. Agrobacterium mediated transformation of Coker 312 hypocotyls was carried out with sgRNA3GhPHYA1::CRISPR / Cas9 cassettes. Somatic embryogenesis has been achieved from the Coker 312 callus cultures harbouring sgRNA3GhPHYA1::CRISPR/Cas9 cassette.

Discerning the Differential Expressed Proteins (DEP) towards leafhopper resistance aids in identifying the key peptides involved in resistance. Unravelled peptides sequence information would help in development of functional markers for marker-assisted selection towards leafhopper resistance in cotton.