Cloning of chit42 gene from Trichoderma atroviride PTCC5220 and its structure analysis

Filamentous fungi such as Trichoderma spp. Produce a variety of chitinase enzymes that degrade chitin and play an important role in biological control of fungal diseases. Chitinases are hydrolysis enzymes which degrade chitin, a linear biopolymer of N-acetylglucosamine, into its component residues by breaking the β-1,4 glycosidic bonds. In this research Trichoderma atroviride PTCC5220 an over producer of chitinase enzyme among 30 Trichoderma sp. isolates used for study of chit42 gene. Two specific primers (CUM1/CDM2) and genomic DNA from T. atroviride were used for chit42 amplification. The amplified DNA fragment (about 1.6 kb) was analyzed and confirmed by restriction pattern using XhoI enzyme.The chit42 DNA fragment was cloned into pUC19 and designated as pMJH1. Comparison of the cloned fragment with the cDNA sequence indicated that this gene contains three short introns (58bp, 62bp and 70 bp long) and also an open reading frame coding for a protein of 421 amino acids. Alignment of the amino acid sequence with other reported Chit42 sequence from Trichoderma sp. (T. aureoviride, T. viride, T. harzianum, T. longibrachiatum and T. koningii) showed 100%, 98%, 98%, 97% and 96% identity, respectively.