Proline and chitosan enhanced efficiency of microspore embryogenesis induction and plantlet regeneration in Brassica napus L.

Abstract

The effects of proline (0, 50, 100, 200 and 500 mg l−1) and chitosan (0, 10, 20, 50 and 100 mg l−1) treatment at three incubation periods (1, 2 and 5 days) were assessed on the efficiency of microsopore embryogenesis induction and subsequently plantlet regeneration in rapeseed (Brassica napus L.) cv. “Hyola 401”. About 30–55 % increase in microspore embryogenesis was observed in the cultures exposed to 100 mg l−1 proline for 2 and 5 days. Higher levels were not advantageous so that, microspore embryogenesis significantly reduced when 200 and 500 mg l−1 proline were exogenously applied to the culture medium. High normal regeneration was achieved in the cultures treated with 50 and 100 mg l−1 proline for 2 and 5 days. In addition, callogenesis increased as proline level and duration of its treatment was increased in which the majority of microspore-derived embryos (78 %) underwent callusing in the cultures exposed to 500 mg l−1 proline for 5 days. In contrast to untreated cultures, the efficiency of microspore embryogenesis increased about 1.8-fold in response to 10 mg l−1 chitosan for 2 days. High levels of chitosan were detrimental to microspore embryogenesis so that embryogenesis was completely inhibited in the cultures exposed to 50 and 100 mg l−1 for 5 days. Chitosan treatment was not advantageous to the normal plantlet development at all levels and durations tested. High levels and durations of chitosan treatment resulted in the higher callusing so that the highest callusing (88 and 79 %) were observed in the presence of 100 mg l−1 for 1 day and 20 mg l−1 for 2 days, respectively. Efficiency of microspore embryogenesis induction and plantlet regeneration could be improved by proline and chitosan treatment when appropriate levels and durations of incubation were selected.