Characterization and comparative analysis of three low molecular weight glutenin C-subunit genes isolated from Aegilops tauschii

Abstract

-M. 2007. Characterization and comparative analysis of three low molecular weight glutenin C-subunit genes isolated from Aegilops tauschii. Can. J. Plant Sci. 87: 273-280. Three low molecular weight glutenin subunit (LMW-GS) genes from T121, T128 and T132 accessions of Aegilops tauschii (DD, 2n = 2x = 14) were amplified using allelic-specific PCR primers. The amplified products with a size of about 900 bp were cloned and sequenced. Three complete coding sequences of LMW-GS with 918 bp, 921 bp and 918 bp were obtained and named as LMW-T121, LMW-T128, LMW-T132, respectively. Each gene contained a complete open reading frame and had no introns. The deduced amino acid sequences showed that all belonged to LMW-m type subunit with a predicted molecular weight of about 32 kDa, corresponding to the size of LMW C-subunits. All three subunits possessed eight cysteine residues and had greater homology with previously characterized LMW-m subunits from bread wheat and related species than LMW-s or LMW-i sequences. Some amino acid substitutions and insertion/deletion variations among the sequences were detected. The corresponding three C-subunits in seed endosperm encoded by LMW-T121, LMW-T128, LMW-T132, respectively, were identified and confirmed by SDS-PAGE, MALDI-TOF-MS and direct N-terminal amino acid sequencing. Phylogenetic analysis demonstrated that LMW-m and LMW-s type subunit genes possessed higher identity and they were obviously separated from LMW-i type subunit genes. The LMW-m type might be the primitive form while the LMW-s and LMW-i types are variant forms.

A Novel Chimeric Low-Molecular-Weight Glutenin Subunit Gene From the Wild Relatives of Wheat Aegilops kotschyi and Ae. juvenalis: Evolution at the Glu-3 Loci

“…With the LMW-GS gene group, three types of LMW-GS genes were divided into different subgroups but LMW-s-and LMW-m-type genes were shown to have higher homology. These results were consistent with their sequence characteristics and recent reports of An et al (2006) and Pei et al (2007). Particularly, the AjkLMW-i gene located on the U genome was clustered in the same subgroup with other LMW-m-type genes and shared 86% homology.…”

“…In the past, an increasing number of LMW-GS genes have been isolated and characterized from common wheat (D'Ovidio and Masci 2004) and high allelic variations have been identified at the Glu-3 locus (Gianibelli et al 2002;Yanet al 2003a,b,c;An et al 2005;Li et al 2006). A recent focus has been to isolate new LMW-GS genes from wheat related species, e.g., Triticum boeoticum (Lee et al 1999), T. monoccocum , Aegilops tauschii ( Johal et al 2004;Pei et al 2007), and T. dicoccoides (Q. Li et al 2008).…”

Molecular cloning and characterization of five novel low molecular weight glutenin subunit genes from Tibetan wheat landraces (Triticum aestivum L.)

“…Genomic DNA was extracted from 3 to 5-day-old seedlings using modified the cetyl trimethyl ammonium bromide (CTAB) procedure (Murray and Thompson 1980).The specific degenerate primers of LMW-GS gene were designed to amplify the coding regions based on previously cloned LMW-GS genes (Ikeda et al 2002;Pei et al 2006). The sequences of two primers were P1: 5 0 -ATGAAGACCTTCCTCG TCTTTG-3 0 , P2: 5 0 -TTATCAGTAGGCACCAACT CC-3 0 (synthesized by Sangong Inc., China).…”

“…The LMW-GS for coding cloned genes were arrowed, the molecular weights of which were approximately 32 kDa. According to former research, MALDI-TOF-MS was the best level of accuracy and resolution when measuring the molecular weights of LMW-GS Pei et al 2006;Feng et al 2011). To obtain accurate molecular weights, LMW-GS was further analyzed by MALDI-TOF-MS. From Table 1 it could be seen that the molecular weights of LMW-1, LMW-2, LMW-3, LMW-4 and LMW-5 by MALDI-TOF-MS were 32,393.36, 32,448.44, 32,518.38, 32,687.13 and 32,483.65 Da, generally consistent with LMW-GS as shown by arrows in Fig.…”

“…Generally, LMW-GS contain eight cysteine residues that are involved in forming intra-and inter-molecular disulfide bonds in the gluten macro polymers (GMP) . In LMW-m and LMW-s type subunits the first cysteine residue is present in the N-terminal domain and the other seven cysteine residues are in the C-terminus, but in LMW-i type subunits all eight cysteine residues are located in the C-terminus (Ikeda et al 2006;An et al 2006;Pei et al 2006).…”