發(fā)布時(shí)間：2018-06-15 11:18

  本文選題：仙茅 + 組織培養(yǎng)�。� 參考：《中南林業(yè)科技大學(xué)》2017年碩士論文

【摘要】：仙茅是我國一味使用了千年的中藥,主要治療陽痿、筋骨萎軟、腰膝冷痹、陽虛冷瀉等癥狀,是非常寶貴的中藥材。野生仙茅因人為和牲畜的破壞,導(dǎo)致數(shù)量急劇減少;但其作為中藥材和保健品原料,市場需求卻越來越大,野生仙茅數(shù)量已經(jīng)無法滿足人們的需要;目前人工方法栽培仙茅已經(jīng)表現(xiàn)出極大的現(xiàn)實(shí)經(jīng)濟(jì)價(jià)值。傳統(tǒng)培育仙茅的方式工作量大,成活率低,且浪費(fèi)了大量優(yōu)質(zhì)仙茅。本論文研究了仙茅更高效、便捷的繁育方式,利用植物組織培養(yǎng)育苗和人工種子技術(shù)育苗,在一定程度上解決了仙茅產(chǎn)量不足的問題,具有較大的現(xiàn)實(shí)意義。本論文來源于國家林業(yè)公益性行業(yè)科研專項(xiàng)項(xiàng)目《仙茅、雞血藤資源培育及林下種植技術(shù)研究》,項(xiàng)目編號(hào)201304807。目前市場上仙茅的品種很多,本研究基于的是栽培于廣州從化的四川宜賓野生仙茅。通過設(shè)置單因素試驗(yàn)、正交試驗(yàn)等探究了快速得到仙茅植株的三條途徑。以仙茅幼葉誘導(dǎo)愈傷組織得到完整植株;由不定芽制作人工種子得到完整植株;利用仙茅的細(xì)胞懸浮培養(yǎng),誘導(dǎo)出胚狀體制作人工種子,以得到完整植株。實(shí)驗(yàn)結(jié)果證明三種方式均具有可行性:正交試驗(yàn)和單因素試驗(yàn)結(jié)果表明,MS基本培養(yǎng)基中加入0.1mg/L的2,4-D、1.5mg/L的KT,3%的多菌靈和8mg/L的檸檬酸,誘導(dǎo)出的仙茅愈傷組織效果最好,可達(dá)96%。方差分析得知2,4-D、KT、多菌靈的濃度對仙茅出愈率的影響差異極顯著,6-BA濃度和NAA濃度對出愈率影響差異極顯著。繼代培養(yǎng)基為MS加入0.3mg/L的NAA,2mg/L的6-BA,0.3mg/L的2,4-D,繼代效果最好,增殖率最高,愈傷組織的質(zhì)量最好,較易誘導(dǎo)出不定芽。在培養(yǎng)基中添加0.3mg/L的NAA有利于生根,移栽至營養(yǎng)土中的成活率可達(dá)到90%以上。在人工胚乳中加入0.05 mg/L6-BA,0.8mg/L NAA制作的人工種子萌發(fā)率可達(dá)到93%,添加3mg/L多菌靈時(shí)人工種子的染菌率最低。對比試驗(yàn)表明包兩層膜的人工種子比包一層膜的人工種子,保水性和機(jī)械性都更好。當(dāng)海藻酸鈉濃度為3%時(shí),制作的人工種子質(zhì)量最好,大小相同,軟硬合適。以2%的CaCl2為固化劑,反應(yīng)兩次,第一次固化5分鐘,第二次固化10分鐘,制成的人工種子形態(tài)狀況最好,顆粒均勻,軟硬適中,呈球形,透明有彈性。愈傷組織繼代時(shí),培養(yǎng)基蔗糖濃度為15g/L,6-BA濃度為2mg/L、NAA濃度為0.5mg/L時(shí),可得到疏松易碎適合做懸浮細(xì)胞的愈傷組織。正交試驗(yàn)探究非激素水平對細(xì)胞懸浮培養(yǎng)影響結(jié)果表明:蔗糖影響懸浮細(xì)胞形態(tài),MES對懸浮培養(yǎng)液的PH值影響較大,對數(shù)據(jù)進(jìn)行方差分析得知,不同濃度的蔗糖、水解酪蛋白、MES、脯氨酸對仙茅細(xì)胞懸浮培養(yǎng)影響的差異極顯著。探究激素對細(xì)胞懸浮培養(yǎng)的影響時(shí)發(fā)現(xiàn),2,4-D影響懸浮細(xì)胞結(jié)構(gòu),當(dāng)2,4-D濃度為0.5mg/L時(shí),胚性細(xì)胞數(shù)量達(dá)到最大值。細(xì)胞的生長隨著培養(yǎng)時(shí)間的變化呈現(xiàn)“S”形曲線,接種量對懸浮培養(yǎng)的影響最大,接種量為15%是仙茅細(xì)胞懸浮培養(yǎng)的最適接種量。本論文以組織培養(yǎng)育苗,替代傳統(tǒng)的根莖繁殖,成功建立了仙茅組織培養(yǎng)繁殖體系,可將其應(yīng)用于仙茅的大規(guī)模生產(chǎn),解決目前仙茅市場緊缺的狀況。并且以此為基礎(chǔ),利用仙茅不定芽,成功研制出仙茅雙層膜人工種子,在一定程度上解決了傳統(tǒng)人工種子抗壓能力弱和保水性能差的問題。成功建立仙茅植物細(xì)胞懸浮體系,為今后在細(xì)胞水平上研究仙茅奠定理論基礎(chǔ)。
[Abstract]:Chinese herb is a Chinese herbal medicine used for thousands of years. It mainly treats impotence, soft bone, cold arthralgia, cold and diarrhea of the waist and knees, Yang deficiency and diarrhea and so on. It is a very valuable Chinese herbal medicine. Wild imfairy is the destruction of people and livestock, resulting in a sharp decrease in quantity. But as a raw material of Chinese medicine and health care products, the demand for the market is getting bigger and bigger, the number of wild imfairy has already been found. It is impossible to meet the needs of people. The cultivation of curculia is of great practical economic value at present. The traditional cultivation method of the Chinese curculus is great work, low survival rate and a large number of high quality curl. This paper studies the more efficient and convenient breeding methods of the grass, the cultivation of seedling and the technology of artificial seed cultivation with plant tissue culture. To a certain extent, it has solved the problem of insufficient yield of curl, which is of great practical significance. This paper comes from the special project of the national forestry public welfare industry, "imimu", the cultivation of the resources of the Caulis, and the research on the planting technology under the forest. The project No. 201304807. is at present a lot of varieties of the green grass on the market. This study is based on the cultivation in Guangzhou. The wild centith in Yibin, Sichuan, Conghua. Through the setting of single factor test and orthogonal experiment, three ways to quickly get the plant of centip were explored. The whole plant was obtained by the induction of the callus from the young leaves of the centian; the artificial seeds were made from the adventitious buds, and the artificial seeds were induced by the suspension culture of the centip. The results of the experiment showed that the three methods were all feasible. The orthogonal test and single factor test showed that the MS basic medium added 0.1mg/L 2,4-D, 1.5mg/L KT, 3% Carbendazim and 8mg/L citric acid, and the induced callus effect was the best, and the 96%. variance analysis showed that 2,4-D, KT, and carbendazim were strong. There are very significant differences in the effect of degree on the recovery rate of curculus. The effect of 6-BA concentration and NAA concentration on the recovery rate is very significant. The subculture medium is MS adding 0.3mg/L NAA, 2mg/L 6-BA, 0.3mg/L is the best, the highest proliferation rate, the best quality of the callus, and the easier induction of adventitious buds. The NAA favorable for adding 0.3mg/L in the medium is favorable. Yu Shenggen, the survival rate of transplanted into the nutrient soil could reach 90%. The germination rate of artificial seeds in artificial endosperm was 0.05 mg/L6-BA, and the germination rate of artificial seeds produced by 0.8mg/L NAA could reach 93%. The rate of infection of artificial seeds was the lowest when adding 3mg/L carbendazim. When the concentration of sodium alginate is 3%, the quality of the artificial seeds is the best, the size is the same, the soft and hard is suitable. With 2% CaCl2 as the curing agent, two times, the first curing 5 minutes, and the second curing 10 minutes, the artificial seeds have the best shape, the particles are uniform, the soft and hard, the transparent and elastic. On the other hand, when the concentration of sucrose is 15g/L, the concentration of 6-BA is 2mg/L and the concentration of NAA is 0.5mg/L, the callus which is loose and suitable for suspension cells can be obtained. The results of the effect of the non hormone level on the cell suspension culture show that the sucrose affects the morphology of the suspension cell, the MES has great influence on the pH value of the suspension culture, and the data are carried out. The variance analysis showed that different concentrations of sucrose, casein hydrolysate, MES, proline had a very significant difference in suspension culture of curculus. When the effect of hormone on cell suspension culture was found, 2,4-D affected the structure of suspension cells. When the concentration of 2,4-D was 0.5mg/L, the number of embryogenic cells reached the maximum. The change showed "S" shape curve, the influence of inoculation on suspension culture is the most, the inoculation amount is 15% is the optimum inoculation quantity of the suspension culture of curculus. This paper successfully established the culture and breeding system of curculus curculera by tissue culture and breeding, instead of the traditional rhizome propagation. On the basis of the market, and using the adventitious bud of the curl, the artificial seed of the bilayer film of the curl grass has been successfully developed. To a certain extent, the problem of the weak compression ability and poor water retention of the traditional artificial seeds is solved. The suspension system of the plant of the plant is successfully established to lay a theoretical foundation for the study of the Curca on the cell level in the future.
【學(xué)位授予單位】：中南林業(yè)科技大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S567.239
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本文編號(hào)：2021828