Cloning and semi-quantitative expression of endochitinase (ech42) gene from Trichoderma spp.
Species of Trichoderma such as T. harzianum, T. viride and T. atroviride are some of the most potent antagonistic fungi, and have been used as plant growth promoters in developed countries. Endochitinase (ech42) gene which is involved in mycoparasitism, was isolated from Trichoderma spp. taken from hot-arid soils of Rajasthan, cloned, sequenced and its expression profiling was carried by reverse transcription-polymerase chain reaction (RT-PCR) technique. The cloned DNA sequence was 1,476 base pairs. Gene encoding endochitinase was ligated in pGEMT cloning vector. The plasmids were transformed in DH5α Escherichia coli competent cells and clones were confirmed through sequencing and restriction analysis. Endochitinase gene expression was then observed for different Trichoderma isolates viz., T. harzianum (T14 and T12) and T. atroviride (T5). Among the three, higher expression of endochitinase was observed in T14 and T12, whereas T5 showed lesser expression with respect to T14 and T12 strain. The Trichoderma chitinase enzyme activity was monitored for all isolates under study. The highest chitinase activity was observed in T14 and T11 viz., 17.21 (1 enzyme μg/ml) and 13.11 enzyme μg/ml, respectively.
Keywords: Endochitinase, cloning, expression, Trichoderma atroviride, Trichoderma harzianum, Trichoderma viride