Induction and evaluation of tetraploids in annual ryegrass

Induced polyploidy by application of mutagens such as colchicine is a well-known technique with a long history. It has been used successfully on many plant species. Generally they have different morphological, physiological, and genetic characteristics than their diploid progenitors. As a result of these differences, they have higher biomass yield than their diploid progenitors as they can be more tolerant to biotic and abiotic stress. Therefore, induced polyploidy is an important breeding method, especially for forage species. Approximately, 50% of the annual ryegrass cultivars registered in the world are autotetraploids although it is a natural diploid with 2n=14 chromosome. Based on the results of previous studies, it has been determined that characteristics and performance of autopolyploids depend greatly on their diploid progenitors. Therefore, it has critical importance to evaluate diploid germplasms in the conditions of target regions and select the best performing ones for poliploidy induction. The objective of this study was to develop an efficient method for our laboratory to induce chromosome doubling in annual ryegrass and develop a tetraploid population. Approximately 1200 seeds of a diploid population developed by 3 cycles of selection from an annual ryegrass collection including 140 gene bank accessions based on their performance in Tekirdağ conditions. Five different doses of colchicine (0.05, 0.1, 0.2, 0.3, 0.4 %) and 3 different period of application (4, 8 and 24 h) were used in the study. Flow cytometry was used in ploidy analysis after 8 weeks of colchicine application. The highest recovery of autotetraploids (15%) was obtained by treatment of one week old germinating seedlings with 0.2% colchicine for 8 hours. A high incidence of mixoploidy was obtained following colchicine treatment. Rarely hexaploids were also encountered. It was found out that ploidy varied among the tillers of the same plants when ploidy analysis repeated after 4 months of colchicine applications. Autotetraploid tillers propagated vegetatively and used in polycrosses to develop an autotetraploid population. After a two-year selection process, the performance of the polyploid population was compared with that of the diploid population and other standards.