Analysis of nucleotide sequences and expression levels of the LEC1 gene in pea varieties with different percentage of seed storage protein

   Peas is one of the best sources of plant protein containing all essential amino acids. Genetic determinants that control biosynthesis and accumulation of seed storage proteins in peas currently remain poorly studied. It is known
that these processes, along with seed development and maturation, are regulated by the genes of transcription factors LEC1, LEC2, ABI3, FUS3.

   In this regard, the purpose of the current study was to analyze the nucleotide sequences of the coding region of the LEC1 gene and its expression levels in high- and low-protein varieties to identify molecular markers associated with protein percentage in pea seeds.

   The experiments were carried out in 2021–2024. The study material was 110 pea varieties and lines from the VIR collection. The percentage of seed storage protein was determined using the Bradford method. DNA was isolated using the Genomic DNA Purification Kit (“Thermo Fisher Scientific”, USA). Primers for the LEC1 gene were first selected using the PrimerSelect program (“DNAStar”, USA). Sequencing was performed on an ABI Prism 3500 genetic analyzer (“Applied Biosystems”, USA). Nucleotide sequences were analyzed using the MegAlign program (“DNAStar”, USA). As a result, there have been selected two groups, including 12 pea varieties and lines with high (23.5 ± 0.4 – 26.1 ± 0.5 %) and 10 with low (18.0 ± 0.3 – 19.7 ± 0.3 %) protein percentage in seeds. Comparative analysis of the nucleotide sequences of the LEC1 gene in high- and low-protein variety samples has allowed identifying previously undescribed single nucleotide substitutions at positions 884 (G/A), 1041 (C/T), 1076 (C/T), 1201 (T/C), 1268 (G/T) bp and an insertion at position 696/697 (C) bp.There has been found that at the early stage of seed development (the 5^th day after pollination) of the most low-protein pea varieties and lines the expression levels of the gene of the transcription factor LEC1 were significantly higher than in the high-protein pea varieties.

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