| The research aiming to improve the system of microspore culture technology was carried out to reveal the influence factors of embryogenesis and plant regeneration in cabbage microspore culture. The system showed high induction frequency and embryoid quality. At the same time, it also could provide much effective process of plant regenation in embryogenesis pathway and callogenesis pathway. In this experiment, main study have been conducted on the embryo induction effect of four different type materials(hybrids, DH lines, inbred lines in early generation, inbred lines in high generation), the donor plants in different overwintering ways, adding different concentrations of NaCl and melatonin on heat shock culture medium; The process of embryoid to regeneration plants has been observed and the effect of melatonin on regenerated plant formation has been investigated.The main conclusions of this research are as follows:1. There were differences of embryoids induction among the 4 types of materials. The material of hybrids has a highest embryoids induction rate of 6.5 embryos / bud. Early generation inbred material has a lowest induction rate of 1.4 embryos / bud. The embryoids of DH materials were observed after culturing 14 d, while the embryoids of high generation inbred material were observed after culturing 27 d.2. The overwintering way of caching in high mountain was conducive to microspore culture of DH131, DH132, E01, and the average frequency of embryoids was up to 1.9 embryo / bud. From single material analysis, the embryoids induction effect of DH132 was better in the plastic greenhouse than caching in high mountain, which shows that the different overwintering way has different effects on embryoids induction in different materials.3. Adding 150~200mg/L NaCl can significantly improve enlargement ratio of microspore, embryos rate, the quality and development of embryoid in cabbage microspore culture.4. Low concentrations of melatonin are beneficial for the embryogenesis in the microspore culture of cabbage. In the microspore culture of E01 and DH132, the enlargement rate and embryo production were highest as 90%, 86% and 5.8 embryo / bud, 7.1 embryo / bud in the concentration of 1.0μmol/mL MEL.5.Plant regeneration can be formed by callogenesis pathway. In the stage of embryo to callus formation, it would experience a short embryoid dormancy period, callus formation period, the stage of callus turnning green, the adventitious buds differentiation period, adventitious bud growth period, adventitious bud rooting period, finally the plant formed.6. Suitable concentration of melatonin treatment can reduce the rate of callus browning, improve callus differentiation of adventitious buds in quantity and quality, and reduce the rate of vitrification shoots. The callus turned green after culturing 1d in the medium with 2.0μmol/mL melatonin. Compared with the control,the browning rate of callus was lowest, decreased 69.4% in the concentration of 2.0μmol/mL MEL. And the number of adventitious buds was most, increased 279.6%; The growth potential of adventitious buds was best in the treatment of 6.0μmol/mL melatonin, and the vitrification rate of buds was lowest, reduced 94.4% than that of CK.7.The suitable concentration of melatonin can accelerate the seedling growth rate, shorten the rooting time and improve the quality of root development. In the treatment of T4, the stems and leaves of adventitious buds grew fastest,and the root systems were most developed; The taproots, lateral roots and fibrous roots were more clear; The number of root tips, root forks and root crossing were more than that of CK. |
