The main seed storage protein in wheat is gluten. It consists of gliadin and glutenins. Gluten gives high elasticity and extensibility during bread making, facilitating the formation of the dough. Rice is the staple food of Sri Lankans but, it has poor dough making ability compared to wheat. The aim of the present work was to characterize, clone and express α-gliadin in the T 0 generation of Bg 250 rice variety as a preliminary step in improving the dough making ability of rice flour. Five α-gliadin recombinant pCR™2.1-TOPO® clones were selected for sequence analysis. Of the five clones, two functional genes and three pseudogenes were identified. Phylogenetic analysis revealed the two functional genes, (accession numbers KC660359 and KC660358) to be closely related to the α-gliadin genes of Triticum monococcum. The α-gliadin gene (KC660359) contained five cysteine residues, one less than the normal occurrence of cysteine residues in α-gliadin genes. To date there are no reports on expression of gliadin gene in transgenic rice. This novel gene was successfully expressed in the Sri Lankan rice variety Bg 250 under the control of the rice GluB-1 endosperm specific promoter.

Glutelins are the primary source of energy storage in the endosperm of rice grains. Among the glutelin promoters, Glutelin B-1 (GluB-1) is widely studied and used in transgenic rice plants to express recombinant proteins in the endosperm. In this study, three regions: 350 bp, 1308 bp and 2300 bp of the GluB-1 promoter were PCR amplified from the genomic DNA of Bg 250 rice variety. The amplified fragments were cloned into pGEM®-T Easy vector for characterisation of GluB-1 promoter. Each region of GluB-1 promoter was separately cloned into the promoterless binary vector pCAMBIA1391Z harbouring the β-glucuronidase (GUS) reporter gene. Putative transgenic plants were generated by Agrobacterium-mediated gene transformation and confirmed by PCR using nopaline synthase terminator primers. All GluB-1 promoter constructs showed expression of the GUS gene in the endosperm of T 0 transgenic plant seeds. The 1308 bp GluB-1 promoter revealed the highest expression as determined by the GUS assay. This indicates the potential of this promoter for expression of recombinant proteins in rice endosperm.

<p class="abstract"><strong>Background:</strong> High molecular weight (HMW) glutenin protein plays a crucial role in determining dough viscoelastic properties that determines the quality of wheat flour. The aim of the present study was to isolate, clone and analyze (<em>in silico</em>) the HMW glutenin gene of <em>Triticum aestivum</em> cultivar Dacke.</p><p class="abstract"><strong>Methods:</strong> Primers were designed to amplify a 2445 bp fragment of HMW glutenin gene. Ax type HMW glutenin gene from <em>Triticum aestivum</em> cultivar Dacke was isolated using PCR and it was sequenced by primer walking.  </p><p class="abstract"><strong>Results:</strong> Amplified HMW glutenin gene was designated as HMWGAx. Sequence analysis revealed a complete open reading frame encoding 815 amino acid residues with N- and C terminal non-repetitive domain and a central repetitive domain. The calculated molecular weight of the deduced HMW glutenin protein was ~88 kDa and the number of cysteine residues in the HMWGAx was four, in accordance with other x type HMW glutenin proteins. Phylogenetic analysis revealed 100% homology to the previously studied Ax2* type HMW glutenin gene from cultivar Cheyenne. Predicted secondary structure results showed that it was similar to1Ax1 type of common wheat (<em>Triticum aestivum</em>), having superior flour milling quality.</p><p><strong>Conclusions:</strong> Sequence analysis suggests that HMWGAx protein significantly and positively correlates with the properties of elasticity and extensibility of gluten. </p>