Somatic Embryogenesis And Morphological And Histological Variations Of Garlic (Allium Sativum L.)

Garlic(Allium sativum L.),is an important medicinal and edible plant.Asexual reproduction by vegetative organs,such as bulbs,is its main reproductive way,but low coefficient of propagation limited mass production of its seedlings.And viral infection caused by long time vegetative propagation may also lead to serious losses in yield and quality.It has been made great progress to obtain virus-free plantlets by tissue culture.However,the propagation coefficient is not high,the domestication and transplanting is difficult,with a high cost,limiting the speed of its promotion.There are two main ways to obtain virus-free plantlets using tissue culture:organogenesis and somatic embryogenesis.Compared with the former,the latter has many advantages,such as high quantity,fast speed,structural integrity and strong genetic stability.Although many progresses have been made in somatic embryogenesis of garlic,the results are not system.Therefore,it is necessary to study the regeneration of garlic plants through somatic embryogenesis deeply.In this paper,the rachis,bulb,root tips and leaf bases of garlic varieties,specifically'Ershuizao','Xuzhou white','Zhengyuezao','Early-maturing'and 'Portuguese garlic'were used as explants to study the influence of different varieties,explants and culture on garlic callus induction and somatic embryogenesis.In addition,morphological and histological observation was carried out to investigate the characteristics of somatic embryogenesis.The main results are as follows:1.There were significant differences in induction time and rate of embryogenic callus among various explants.The rachi has the quickest induction time of embryogenic callus,averaged 9 days,and the induction rate was approximately 90.18%.The trend of callus formation time and induction rate among different garlic varieties was similar.Among different varieties,'Ershuizao' and 'Xuzhou white' are better than 'Zhengyuezao','Early-maturing' and 'Portuguese garlic'.2.The optimum medium for induction of embryogenic callus was B5+2,4-D 2.5-3.5 mg·L-1+KT 0.5-1.0 mg·L-1+sucrose 30-40 g·L-1+agar 6.5 g·L-1,pH5.8-6.2.Compared with the solid culture,the liquid suspension culture has higher callus proliferation rate.In the medium of 6-BA 1.0 mg·L-1 + KT 0.5 mg·L-1,or 6-BA 1.5 mg·L-1 +NAA 0.1 mg·L-1,the percentage of weight gain of callus reached about 44%.The medium 6-BA 2.0 mg·L-1+KT 1.0 mg·L-1 has the highest somatic embryo germination rate.3.The morphology of callus induced by various explants and garlic varieties was different.The rachis of 'Ershuizao' has formed the largest number of embryogenic callus,and the callus had a good quality with consistent size,smooth surface and easily dispersed oval-shaped granules,so the rachis of 'Ershuizao,was a good regeneration material for the efficient induction of somatic embryogenesis of garlic.Morphology and Histological observation of different developmental stages revealed that somatic embryogenesis was similar to that of the zygotic embryo.The embryonic callus was formed by dedifferentiation from explants surface parenchyma cell,which had bigger nuclei,thick cytoplasm,clear nucleoli and arranged densely.after,globular embryo.And the globular embryo transformed into the shield embryo from the ends.At this time,big gap formed around the embryo.Bud and root primordium were then formed from the ends of the shield embryo,and mature somatic embryo formed.Then the mature embryos turned green at the center of somatic embryos.Roots and shoots germinated from the greening point to form plantlet.But suspension culture was easy to form non-embryonic cells with irregular sharp,bulky cells,small nuclear,scarce cytoplasm,shallow staining,and the cell metabolism was weak and the differentiative capacity was poor.