Molecular characterization and SNP identification using genotyping-by-sequencing in high-yielding mutants of proso millet

Neethu Francis; Ravikesavan Rajasekaran; Veera Ranjani Rajagopalan; S Vinothini Bakya; Raveendran Muthurajan; Ashwini Girish Kumar; Senthil Alagarswamy; Iyanar Krishnamoorthy; Chitdeshwari Thiyagarajan

doi:10.3389/fpls.2023.1108203

Molecular characterization and SNP identification using genotyping-by-sequencing in high-yielding mutants of proso millet

Front Plant Sci. 2023 May 18:14:1108203. doi: 10.3389/fpls.2023.1108203. eCollection 2023.

Authors

Neethu Francis¹, Ravikesavan Rajasekaran², Veera Ranjani Rajagopalan³, S Vinothini Bakya¹, Raveendran Muthurajan⁴, Ashwini Girish Kumar⁵, Senthil Alagarswamy⁶, Iyanar Krishnamoorthy⁷, Chitdeshwari Thiyagarajan⁸

Affiliations

¹ Department of Genetics and Plant Breeding, Centre for Plant Breeding and Genetics, Tamil Nadu Agricultural University, Coimbatore, India.
² Centre for Plant Breeding and Genetics, Tamil Nadu Agricultural University, Coimbatore, India.
³ Department of Plant Biotechnology, Centre for Plant Molecular Biology and Biotechnology, Tamil Nadu Agricultural University, Coimbatore, India.
⁴ Directorate of Research, Tamil Nadu Agricultural University, Coimbatore, India.
⁵ Parse Biosciences, Stockholm, Sweden.
⁶ Department of Crop Physiology, Tamil Nadu Agricultural University, Coimbatore, India.
⁷ Department of Millets, Centre for Plant Breeding and Genetics, Tamil Nadu Agricultural University, Coimbatore, India.
⁸ Department of Soil Science and Agricultural Chemistry, Tamil Nadu Agricultural University, Coimbatore, India.

Abstract

Proso millet (Panicummiliaceum L.) is a short-duration C₄ crop that is drought tolerant and nutritionally rich and can grow well in marginal lands. Though the crop has many climate-resilient traits like tolerance to drought and heat, its yield is lower than that of common cereals like rice, wheat, and maize. Being an underutilized crop, the molecular resources in the crop are limited. The main aim of the present study was to develop and characterize contrasting mutants for yield and generate functional genomic information for the trait in proso millet. Gamma irradiation-induced mutant population was screened to identify high-yielding mutants, which were evaluated up to M₄ generation. One mutant with a dense panicle and high yield (ATL_hy) and one with a lax panicle and low yield (ATL_ly) along with the wild type were sequenced using the genotyping-by-sequencing approach. The variants detected as single nucleotide polymorphisms (SNPs) and insertions-deletions (InDels) were annotated against the reference genome of proso millet. Bioinformatic analyses using the National Center for Biotechnology Information (NCBI) and UniProt databases were performed to elucidate genetic information related to the SNP variations. A total of 25,901, 30,335, and 31,488 SNPs, respectively, were detected in the wild type, ATL_hy mutants, and ATL_ly mutants. The total number of functional SNPs identified in high-yielding and low-yielding mutants was 84 and 171, respectively. Two functional SNPs in the high-yielding mutant (ATL_hy) and one in the low-yielding mutant (ATL_ly) corresponded to the gene coding for "E3 ubiquitin-protein ligase UPL7". Pathway mapping of the functional SNPs identified that two SNPs in ATL_ly were involved in the starch biosynthetic pathway coding for the starch synthase enzyme. This information can be further used in identifying genes responsible for various metabolic processes in proso millet and in designing useful genetic markers.

Keywords: SNP; genotype-by-sequencing; high yield; molecular; mutants; proso millet.