Allelic Variation in Zmfatb Gene Defines Variability for Fatty Acids Composition Among Diverse Maize Genotypes

Abstract

Edible oil with lower saturated fatty acids is desired for perceived quality and health benefits to humans and livestock. fatb gene encoding acyl-ACP thioesterase is a key player in the conversion of palmitic acid to oleic acid, thereby modifying the ratio of saturated to unsaturated fatty acids in maize kernels. The present investigation characterised the full-length sequence of the Zmfatb gene (4.63 kb) in two mutants (Zmfatb) and eight wild-types (ZmfatB) inbreds to study allelic variation, gene-based diversity, phylogenetic-relationship, protein-modelling, and molecular-docking to identify novel candidates for modification of fatty acid profile. Sequence alignment revealed wide genomic variability for Zmfatb among the inbreds; identified five novel SNPs and two InDels that clearly differentiated the wild-type and mutant genotypes. Gene-based diversity using 11-InDel markers categorised 48-diverse maize-inbreds into two-clusters. The majority of mutant and wild-type inbreds were grouped in separate clusters and led to the generation of 41 haplotypes. Genetic relationship of maize fatb gene with orthologues among 40 accessions of 12 oilseed-crops using both nucleotide and protein sequence clustered maize, soybean, sunflower, opium-poppy, Citrulus lanata, quinoa, and prunus species into one cluster; and brassica, camelina, and arabidopsis into the different cluster. The clustering pattern revealed that the plant oil with higher unsaturated fatty acids, particularly oleic, linoleic, and linolenic acids grouped together in one cluster and higher proportions of other fractions like arachidic, eicosenoic, and erucic acids grouped in another cluster. Physico-chemical properties highlighted more similarity between maize and 29 orthologue proteins, but orthologues were found to have better thermostability. Homology models have been developed for maize mutant and wild-type inbreds using Umbellularia californica (PDB ID: 5x04) as a template. Predicted protein models possessed optimum confidence-score and RMSD values and validated stability via., Ramachandran plots. Molecular docking indicated most of the interactions of protein-ligand were having similar binding-affinity due to the broader specificity of fatty acyl-ACP thioesterases and the presence of conserved-domains across crops. This is the first report on the comprehensive molecular characterisation of the fatb gene in maize and various orthologues. The information generated here provided new insights into the genetic diversity of fatb gene which can be utilised for the enhanced nutritive value of oil in the breeding programme.

Molecular Characterization and Haplotype Analysis of Low Phytic Acid-1 (lpa1) Gene Governing Accumulation of Kernel Phytic Acid in Subtropically-Adapted Maize

“…In comparison to Hossain et al [54] (1.81 alleles/loci), Chhabra et al [49] (2.00 alleles/loci) and Katral et al [52] (2.27 alleles/loci), the average number of alleles per locus detected in the present investigation was found to be higher (2.53 alleles/loci). The present research reported lower mean heterozygosity, which showed that most of the loci were homozygous and the alleles were stable [52]. Of the 48 genotypes examined in the study formed 42 haplotypes.…”

“…The reference sequence showed more similarity with mutants viz., PMI-LP1-124 and A632 lpa1-1. The evolutionary distance is regarded to be a powerful predictor of the neutral mode of evolution based on synonymous (Ks) and nonsynonymous (Ka) sites [52]. The direction and strength of natural selection operating on protein-coding genes is determined from the Ka/Ks ratio and a ratio of <1 denotes stabilizing selection [53].…”

“…Gene-based markers used in genetic diversity analysis provide opportunity to evaluate genetic relatedness and locus conservation within genes with greater efficiency [52]. In comparison to Hossain et al [54] (1.81 alleles/loci), Chhabra et al [49] (2.00 alleles/loci) and Katral et al [52] (2.27 alleles/loci), the average number of alleles per locus detected in the present investigation was found to be higher (2.53 alleles/loci). The present research reported lower mean heterozygosity, which showed that most of the loci were homozygous and the alleles were stable [52].…”

Allelic variation in shrunken2 gene affecting kernel sweetness in exotic-and indigenous-maize inbreds

“…InDel markers are co-dominant and involves simple and easy PCR assay, and requires standard gel electrophoresis for separation of amplicons. Further, InDel -based assay does not involve high cost as required in sequence-based approach, as shown in similar studies carried out on su1 and fatb genes for allelic diversity and comprehensive molecular characterization [ 2 , 57 ]. Our study showed that Sh2 -specific InDel markers developed in the present study would be useful in identifying haplotypes of unknown lines to understand their lineage.…”

Multi‐omics‐driven advances in the understanding of triacylglycerol biosynthesis in oil seeds

“…Furthermore, a G‐base insertion in the sixth exon of the FATB allele led to a premature stop codon and lower levels of SFAs in maize (Zheng et al., 2014). Sequence alignment of ZmFATB in several maize germplasms revealed extensive genetic variation and identified five novel SNPs and two InDels (Katral et al., 2022). The SFAs levels were substantially reduced in CRISPR/Cas9‐mediated knockout plants for the downregulation of the FATB1a or FATB1b gene in soybean (Ma et al., 2021).…”