Functional screening yields a new beta-1,4-endoglucanase gene from Heterodera glycines that may be the product of recent gene duplication

Clones with secreted cellulolytic activity were identified when a cDNA libr ary constructed from poly A(+) RNA of preparasitic second-stage juveniles o f Heterodera glycines, the soybean cyst nematode, was expressed in the Esch erichia coli SOLR strain and overlaid with a carboxymethylcellulose (CMC) s ubstrate. Twenty CMC-degrading clones were analyzed, and all were either id entical or strongly similar to a beta -1,4-endoglucanase gene (HG-eng-2), p reviously isolated from H, glycines. A subgroup of identical "HG-eng-2-like " clones had considerable differences in the 5' untranslated region compare d with HG-eng-2 and were designated HG-eng-3, One H, glycines genomic done contained HG-eng-2 and HG-eng-3 full-length genes, separated by a distance of approximately 8 kb, and a second genomic clone contained two copies of H G-eng-2, separated by approximately 6.5 kb, suggesting the presence of endo glucanase gene clusters in H, glycines. The HG-eng-2 and HG-eng-3 genes wer e in opposite transcriptional orientation, with considerable nucleotide dif ferences in their 5' flanking regions. The highly conserved nucleotide sequ ence in the introns and exons and their close proximity within the genome s uggest that HG-eng-2 and HG-eng-3 are the products of recent gene duplicati on and inversion.