ABSTRACT In order to study the effect of bio-fertilizers, micronutrients and N mineral application on barley plant under calcareous soil conditions. Field experiment was conducted at Experimental Research Station –Ras Sudr, D.R.C. in 2017-and 2018 winter seasons. Experimental factors were: micronutrients as foliar application (Fe, Mn Zn at 250 ppm), mineral N (50, 75 and100 kg N/fed) and Bio-fertilizers (control, Azotobacter chroococcum and Azospirillum brasilense either single or mixed application). The obtained results  found that, significant effects of either bio-fertilizers, mineral N and micronutrients as foliar application on yield parameters, mineral contents of barley plants, yield and its attributes, P and N mineral contents of barley grains and microbial activity in barley plant rhizosphere. Interaction of mixed bio-fertilizers treatment micronutrients foliar application of 250 ppm gave maximum enhancement for most studied treatments.  Barley straw and grains yield during the two successive seasons. While P, N contents of both barley shoots and grains yield recorded maximum values with mixed bio-fertilizers treatment, mineral N and Micronutrients as foliar application of 250 ppm. The most effect treatment for microbial counts of bio-fertilizers, yield parameters and nutrients content was Mixture bio-fertilizers Mineral N at 100 units/fed and Mocronutrients 250ppm.