‘宁杞8号'体细胞胚胎发生体系建立

Establishment of somatic embryogenesis system of Lycium barbarum ‘Ningqi 8'

关键词:  ‘宁杞8号', 体细胞胚胎发生, 体胚诱导增殖, 体胚萌发, 植株再生

Key words:  L. barbarum ‘Ningqi-8', somatic embryogenesis, somatic embryo induction and proliferation, somatic embryo germination, plant regeneration

摘要:

以宁夏枸杞新品种‘宁杞8号'幼嫩叶片为外植体, 探讨激素组合及添加物对‘宁杞8号'体细胞胚胎诱导、体胚增殖、萌发和植株再生的影响,并建立高效稳定的体细胞胚胎发生体系。结果表明:通过对6-BA、2,4-D和IAA进行正交分析,筛选出‘宁杞8号'体细胞胚诱导最优激素组合:6-BA 1.0 mg·L^-1+2,4-D 0.3 mg·L^-1 + IAA 0.4 mg·L^-1,体胚诱导率达88.67%。极差分析比较各激素间的影响,6-BA对体胚诱导影响最显著。当高浓度的生长素及低浓度细胞分裂素浓度适宜的配比时,才能诱导产生形态正常数量多的‘宁杞8号'体细胞胚。在体胚增殖培养中,6-BA的浓度过高易导致玻璃化,不利于‘宁杞8号'体胚增殖生长。随着激素浓度的增高,体胚增殖倍数增加,玻璃化率也越高,综合分析得到‘宁杞8号'最佳体胚增殖培养基为6-BA 0.4 mg·L^-1+NAA 0.6 mg·L^-1。当添加IBA 0.3 mg·L^-1+GA₃ 0.4 mg·L^-1+蔗糖10 g·L^-1时,‘宁杞8号'体胚萌发率最高,萌发率达89.17%。添加GA₃及低浓度蔗糖能促进成熟的体胚萌发。对‘宁杞8号'体萌发的影响程度依次是IBA> 蔗糖 > GA₃。活性炭能有效提高‘宁杞8号'体胚再生植株率,同时对萌发体胚的根的发育也有促进作用,当IBA 0.1 mg·L^-1+KT 0.4 mg·L^-1+活性炭1 g·L^-1时,‘宁杞8号'体胚再生植株效果最佳,体胚再生率达91.67%

Abstract:

In order to establish a high-frequency regeneration system for Lycium barbarum. The leaf of L. barbarum ‘Ningqi-8' was selected as material to explore the crucial effects of hormone combinations and additives on the induction of somatic embryogenesis, somatic embryo proliferation and germination and plant regeneration. The results showed that by orthogonal analysis of 6-BA, 2,4-D and IAA, the optimal hormone combination of L. barbarum ‘Ningqi-8' somatic embryos was 6-BA 1.0 mg·L^-1+2,4-D 0.3 mg·L^-1+IAA 0.4 mg·L^-1, the highest induction rate could get to 88.67%. Range analysis showed that 6-BA had the most significant effect on somatic embryogenesis. Using suitable proportion of hormone combination was possible to induce somatic embryogenesis of L. barbarum ‘Ningqi-8', which had a large number of normal morphology. In the culture of somatic embryos, the high concentration of 6-BA was easy to cause vitrification, which was not conducive to the growth and proliferation of L. barbarum ‘Ningqi-8' somatic embryos. The proliferation rate of somatic embryos increased with the increase of hormone concentration, and the vitrification rate increased, too. Then the most efficient proliferation medium for L. barbarum ‘Ningqi-8' was 6-BA 0.4 mg·L^-1+NAA 0.6 mg·L^-1, and the highest germination rate of L. barbarum ‘Ningqi-8' somatic embryo was obtained when IBA 0.3 mg·L^-1+GA₃ 0.4 mg·L^-1+ sucrose 10 g·L^-1 was added. The highest germination rate could get to 89.17%, and the degree of influence on the germination of L. barbarum ‘Ningqi-8' was IBA > sucrose > GA₃. Adding GA₃ and low concentration sucrose could promote the mature somatic embryo germination. Activated carbon could effectively improve the regeneration rate of L. barbarum ‘Ningqi-8' somatic embryo, and simultaneously promote the development of the root of germination embryo. IBA 0.1 mg·L^-1+KT 0.4 mg·L^-1+ activated carbon 1 g·L^-1 was found to be the suitable medium for regeneration in which L. barbarum ‘Ningqi-8' demonstrated the highest multiplying capacity, and the regeneration rate could get to 91.67%.