Random amplification of genomic ends (RAGE) as an efficient method for isolation and cloning of promoters and uncloned genomic regions

Abstract

Isolation of complete coding sequences and regulatory regions is critical for the complete characterization of a gene. Efficient methods to obtain complete genomic or regulatory is important in the process of isolation. The utility of the available genome walking methods are influenced by factors like the size of the genome and the length of the desired sequence. This study utilizes a genome walking method - random amplification of genomic ends (RAGE) efficiently to obtain the 5' – regulatory sequence of a rice stress inducible gene OsAsr1 and to obtain the full length sequence and promoter of the HetR gene of Cylindrospermum stagnale (Cylindrospermum sp. A1345). We demonstrate that this technique can be used for cloning of full length gene and promoters in organisms where whole genome data is unavailable utilising very little sequence information. Our studies show that RAGE can be a strong tool in functional genomics especially in the study of promoters.