Molecular Cloning and Characterization of Novel Phytocystatin Gene from Turmeric,Curcuma longa

Abstract

Phytocystatin, a type of protease inhibitor (PI), plays major roles in plant defense mechanisms and has been reported to show antipathogenic properties and plant stress tolerance. Recombinant plant PIs are gaining popularity as potential candidates in engineering of crop protection and in synthesizing medicine. It is therefore crucial to identify PI from novel sources like Curcuma longa as it is more effective in combating against pathogens due to its novelty. In this study, a novel cDNA fragment encoding phytocystatin was isolated using degenerate PCR primers, designed from consensus regions of phytocystatin from other plant species. A full-length cDNA of the phytocystatin gene, designated CypCl, was acquired using 5′/3′ rapid amplification of cDNA ends method and it has been deposited in NCBI database (accession number KF545954.1). It has a 687 bp long open reading frame (ORF) which encodes 228 amino acids. BLAST result indicated that CypCl is similar to cystatin protease inhibitor from Cucumis sativus with 74% max identity. Sequence analysis showed that CypCl contains most of the motifs found in a cystatin, including a G residue, LARFAV-, QxVxG sequence, PW dipeptide, and SNSL sequence at C-terminal extension. Phylogenetic studies also showed that CypCl is related to phytocystatin from Elaeis guineensis.

Programmed knockout mutation of liver fluke granulin attenuates virulence of infection-induced hepatobiliary morbidity

“…Genomic DNA was extracted from individual adult flukes using the QIAamp DNA Mini Kit (Qiagen). A dual RNA and DNA extraction was used for individual worms at day 60 after infection of hamsters with Ov- GRN - 1 gene-edited NEJ, using RNAzol RT and DNAzol (Molecular Research Center, Inc.) (Chan et al, 2014; Chen et al, 2010). In brief, each worm was homogenized in RNAzol RT using a motorized pestle, the DNA and protein from the lysate was precipitated using DNAse-RNAse-free water.…”

Purification and biochemical characterization of phytocystatin fromBrassica alba

“…Till date, Phytocystatins have been isolated and characterized from diverse sources such as Turmeric (Chan et al, 2014), apple (Ryan et al, 1998), strawberry (Martinez et al, 2005), rice (Abe et al, 1987), Phaseolus mungo and pine apple (Shyu et al, 2004).…”

Review: The future of cystatin engineering

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