Genotype characterizations of some isolated Azospirillum spp. of wheat and maize fields by using rep-PCR

Azospirillum comprises rhizobacteria that live freely in the soil where they change gaseous nitrogen to nitrite and nitrate and as a result promote growth and yield of some economically important plants. The aim of this study was to isolate and identify different native species of Azospirillum and to investigate their genetic diversity as plant growth promoting factors.

In this research, some soil and root samples from wheat and maize fields of Isfahan province were collected and cultured in the tubes containing nitrogen free bromothymol blue (NFB) semisolid media. The Azospirillum isolates were chosen on the basis of morphological properties on NFB and RC (Rojo Congo) media and then were purified. The detection and identification of Azospirillum isolates was done by molecular tests using PCR (Polymerase Chain Reaction), 16S rRNA gene sequencing and genomic fingerprinting with rep-PCR (repetitive extragenic palindromic elements-polymerase chain reaction).

The replication of 646 and 263 bp fragments by Az16S-A, Azo494-F/Azo756-R primers respectively, confirmed the presence of Azospirilum bacteria. Azospirillum species were also identified as A. brasilense, A. lipoferum and A. zeae using replication and sequencing of 16S rRNA gene. The genomic profilings of different Azospirillum isolates produced unique band patterns through application of repetitive element BOX (BOX-AIR), ERIC (ERIC IR, ERIC 2: Enterobacterial Repetitive Intergenic Consensus) and REP (REP IR, REP 2: Repetitive Extragenic Palindromic) and showed high genetic diversity between the isolates.