Computational Insight into PCR Amplified Pectate Lyase Genes from Different Species of Aspergillus
Manish Kumar
Department of Biotechnology, D. D. U. Gorakhpur University, Gorakhpur (U.P.) - 273 009, India
Jeya Nasim
Department of Biotechnology, D. D. U. Gorakhpur University, Gorakhpur (U.P.) - 273 009, India
Sangeeta Yadav
Department of Biotechnology, D. D. U. Gorakhpur University, Gorakhpur (U.P.) - 273 009, India
Aiman Tanveer
Department of Biotechnology, D. D. U. Gorakhpur University, Gorakhpur (U.P.) - 273 009, India
Dinesh Yadav *
Department of Biotechnology, D. D. U. Gorakhpur University, Gorakhpur (U.P.) - 273 009, India
*Author to whom correspondence should be addressed.
Abstract
Pectate lyase (PL) is an important member of pectinase group of enzyme associated with cleaving α -1, 4 linkages in pectate polymers by a β-elimination mechanism, producing 4, 5–unsaturated oligogalacturonates. In the present study, PCR amplification of pectate lyase genes from different species of Aspergillus (namely A. oryzae, A. flavus, A. terreus, A. fumigatus and A. niger) was attempted. The expected size bands resulting from amplification using different sets of primers were gel eluted and sequenced using gene specific primers. The sequences of putative 23 pectate lyase (PL) genes subjected to BLAST analysis revealed its identity to pectate lyase sequences available in databases. Ten sequences of PCR amplified PL genes of A. oryzae namely PL-001 to PL-010 representing five identified PL genes from two different strains revealed maximum identity with the available sequences of A. oryzae RIB40 pectate lyases, A. flavus NRRL 3357 in the range of 98-100%. The phylogenetic tree constructed based on protein sequence of putative PL genes representing different strains of A. oryzae, A. terreus, A. flavus, A. fumigatus revealed several clusters and sub-clusters. A total of 12 PL gene sequences representing A. oryzae MTCC 3782, A. oryzae MTCC 6993, A. fumigatus MTCC 3070, A. fumigatus MTCC 2584 and Aspergillus flavus MTCC 8835 were submitted to GenBank and were assigned accession numbers KP869835 to KP869846.
Keywords: Pectate lyase, PCR amplification, multiple sequence alignment, motif, BLAST, Aspergillus