Organization and structure of NADH-dependent glutamate synthase gene from rice plants

Abstract

Genomic clones for NADH-dependent glutamate synthase (NADH-GOGAT; EC 1.4.1.14) were obtained from a genomic library of rice ( Oryza sativa L. cv. Sasanishki). A genomic clone (λOS42, 14 kb) covered an entire structural gene and a 3.7 kb 5′-upstream region from the first methionine. Another clone (λOS23, 14 kb) contained a 2.8 kb 3′-downstream region from the stop codon. A 7047 bp long clone (λOSR51) consisting of full length cDNA for NADH-GOGAT was isolated from a cDNA library prepared using mRNA from roots of rice seedlings treated with 1 mM NH 4Cl for 12 h. The presumed transcribed region (11.7 kb) consisted of 23 exons separated by 22 introns. Rice NADH-GOGAT is synthesized as a 2166 amino acid protein with a molecular mass of 236.7 kDa that includes a 99 amino acid presequence. DNA gel blot analysis suggested that NADH-GOGAT occurred as a single gene in rice. Primer extension experiments map the transcription start of NADH-GOGAT to identical positions. The 3.7 kb 5′-upstream region was able to transiently express a reporter gene in cultured rice cells. Putative motifs related to the regulation of NADH-GOGAT gene expression were looked for within the 5′-upstream region by database.