枸杞体细胞胚发生过程中Ag+对痕量金属离子吸收的影响

以药用植物宁夏枸杞愈伤组织为材料,离体培养诱导体细胞胚发生.采用多重示踪剂和γ射线能谱分析法研究不同浓度AgNO     

枸杞体细胞胚发生过程中Ag~ 对痕量金属离子吸收的影响

以药用植物宁夏枸杞愈伤组织为材料,离体培养诱导体细胞胚发生。采用多重示踪剂和γ射线能谱分析法研究不同浓度AgNO_3处理的枸杞体细胞胚发生过程中对多种痕量金属元素离子的吸收。结果表明:(1)当AgNO_3的浓度小于50mg/L时,随着AgNO_3浓度的增加,多种痕量金属离子的吸收率也随之增加,而超过此浓度后,对多种痕量金属离子的吸收影响不同。Ag~ 对痕量金属离子的吸收有协同,拮抗或竞争的作用。(2)适当浓度的AgNO_3对细胞分化及体细胞胚发生有促进作用。当AgNO_3的浓度小于50mg/L时,随着AgNO_3浓度的增加,体细胞胚的发生频率随之增加。Ag~ 对枸杞体细胞胚发生表现促进作用,当AgNO_3的浓度为50mg/L时,可大大提高愈伤组织中体细胞胚发生,是对照(不加AgNO_3)组的3倍左右。而超过此浓度后,Ag~ 对枸杞体细胞胚发生表现毒害作用,体细胞胚的发生受到明显抑制。     

枸杞体细胞胚发生过程中Ag^＋对痕量金属离子吸收的影响

We used callus of medical plant, Ningxia Lycium barbarum. L as material to induce somatic embryo genesis, and used multitracer technique and gamma-ray energy spectrum analytical method to study different concentration of AgNO3 on some absorption of trace metal ions during the somatic embryogenesis of Lycium barbarum. L. The results show that (1) when the concentration of AgNO3 is less then 50 mg/L, with the increase of AgNO3 concentration, the absorption rate of some metal ions is increasing, but when the concentration is excess of such concentration, the effects on the absorption rate are different. Ag+ has the effect of coordination, resistance or competition on the absorption of metal ions. (2) AgNO3 in suitable concentration will promote cell differentiation and somatic embryogenesis. When the concentration of AgNO3 is less then 50 mg/L, along with the increase of the concentration, the frequency of somatic embryogenesis is increased, and Ag+ has the promotion on the somatic embryogenesis of Lycium barbarum. L. While the concentration is up to 50 mg/L, it will mostly raise somatic embryogenesis in calli, the amount of callus is about 3 times as much as that of contrast group (in which AgNO3 is not added). When excess of this concentration point, Ag+ has toxic effect, and the somatic embryogenesis is inhibited remarkably.     

宁夏枸杞体细胞胚发生过程中对金属离子的吸收速率和游离氨基酸含量的变化

采用多重示踪技术研究宁夏枸杞 (Lyciumbar barumL .)体细胞胚发生中对多种金属离子的吸收动态及其与游离氨基酸含量变化之间的关系。结果表明 :(1)在枸杞体细胞胚发生中对一些金属离子具有选择吸收特性 ,而且在体细胞胚发育不同时期对同一种金属离子的吸收量也不同 ;(2 )在枸杞体细胞胚发生早期对多数金属离子吸收量迅速增加 ,而后下降。到球形胚期吸收量达到第二个峰值 ,而且金属离子被吸收后提高了体细胞胚发生的频率 ;(3)枸杞体细胞胚发生中游离氨基酸总量从胚性细胞启动期开始下降 ,到胚性细胞形成期到达谷底 ,然后开始上升 ,到多细胞原胚期达到峰值 ,多数游离氨基酸含量变化与金属离子被吸收的量相交叉 ;(4 )外加RbCl和SrCl2 对枸杞体细胞胚发生具有促进作用 ,而且加大了几种游离氨基酸含量变化的幅度。文章讨论了它们之间的关系及其可能的作用机理     

枸杞体细胞胚发生过程中内源多胺代谢动态的研究

完全脱分化的枸杞继代愈伤组织在转入分化培养后第1天就开始启动分化,接着体细胞转变为胚性细胞,继而分裂形成多细胞原胚,球形胚和成熟胚等。与此同时,Put含量迅速上升形成第1个峰值,随后有所下降,但到多细胞原胚期Put含量又上升,并达到最高峰,为对照的6倍,Spd只在胚性细胞分化早期出现,Spm仅在体细胞胚发育晚期存在,外源Put不仅可提高体细胞胚发生频率,而且使3种内源多胺含量均有所提高,加入多胺生物合成抑制剂DFMA后,多胺水平下降,体细胞胚发生几乎完全被抑制。补充外源Put后,多胺的生物合成得到部分恢复,DFMA对体细胞胚发生的抑制效应也被部分解除。结果表明,维持一定量的多胺水平是枸杞体细胞胚发生的必要因素。     

枸杞体细胞胚发生中DNA代谢动态的立体计量

本文以宁夏枸杞无菌苗叶片为材料,离体培养,并诱导体细胞胚胎发生。根据细胞形态计量学原理,应用数学图像处理软件计量由光学底片经Ａ／Ｄ转换成的数学图像中的ＤＮＡ大分子,对枸杞体细胞胚发生过程中ＤＮＡ分子的代谢动态进行量化分析。结果表明：在整个体细胞胚发和过程中ＤＮＡ代谢呈现动态变化。非胚性细胞与胚性细胞期的量化值分别为１．８２％和１．９１％;在二细胞胚、四细胞胚、多细胞胚时期ＤＮＡ缓慢增长,随着胚性愈     

枸杞体细胞胚发生中蛋白质代谢动态的立体计量

以宁夏枸杞无菌苗叶片为材料,离体培养并诱导体细胞胚胎发生。根据细胞形态计量学原理,应用数字图像处理软件计量由光学底片经Ａ／Ｄ转换成的数字图像中的蛋白质大分子,对于枸杞体细胞胚发生过程中蛋白质分子的代谢动态进行了量化处理,并对量化结构分析了蛋白质代谢动态与体细胞胚发生、发育的关系。     

ABA对枸杞体细胞胚发生的调节作用

Using Enzyme Linked Immunosorbent Assay (ELISA) method, we determined the ABA contents of different stages in somatic embryogenesis. The results showed that endogenous ABA contents increased to maximum value twice during somatic embryogenesis. After first maximum value of ABA contents embryogenic cells were observed in callus, and simultaneously, there was a specific protein of somatic embryogenesis investigated by SDS-PAGE. This protein accumulates preferentially in embryogenic callus but not in transferred callus. So it is suggested that ABA could promote the expression of specific genes and the synthesis of embryogenic protein during somatic embryogenesis in Lycium barbarum L. and ABA play an important role in globular stage as well. In addition, treatment of non-embryogenic activity callus with 4 mumol/L exogenous ABA could stimulate somatic embryogenesis. And the ABA function mechanism in relation to somatic embryogenesis was discussed.     

枸杞体细胞胚发生中DNA代谢动态的立体计量

本文以宁夏枸杞无菌苗叶片为材料,离体培养,并诱导体细胞胚胎发生。根据细胞形态计量学原理,应用数字图像处理软件计量由光学底片经A/D转换成的数字图像中的DNA大分子,对枸杞体细胞胚发生过程中DNA分子的代谢动态进行量化分析。结果表明：在整个体细胞胚发生过程中DNA代谢呈现动态变化。非胚性细胞与胚性细胞期的量化值分别为1.82%和1.91%;在二细胞胚、四细胞胚、多细胞胚时期DNA缓慢增长,随着胚性愈伤组织的发育,DNA的含量在梨形胚时期达到高峰;成熟胚的DNA含量虽有所下降,但仍维持较高水平。因此DNA的合成动态变化与体胚生长发育和细胞增殖密切相关。     

Uptake Rate of Tracer MetalIons by a?Lycium barbaruma?Somatic Embryogenesis

The callus of medical plant Lycium barbarum L. in Ningxia was used as the materials for inducing somatic embryogenesis, and multitracer technique and γ-ray energy spectrum analysis were used for studying the rates of uptake of several tracer metal ions during embryogenesis. The results were: 1) Of 23 radionuclides, only 16 (Mn-52, Zn-65, Co-56, Co-60, As-74, Se-75, mTc-95, Rh-99, V-48, Rb-83, Rb-84, Sr-85, Y-87, Y-88, Zr-88 and Zr-89) were selectively absorbed, the other 7 (Be-7, Tc-96, Ru-97, mRh-101, Sc-46, Ga-67 and Cr-51) were not absorbed. 2) The rates of absorption of different metal ions were different at the same developmental stage. In 10 day culture, the rates of uptake of most metal ions were 5%/g, but that of Se-75, Ab-83, Sr-85 and Zr-89 were 10%-30%/g. The rates of absorption of the same metal ion were different at the same developmental stage, e.g. the rates of uptake of Se-75 were increasing in the course of development, two peaks of absorption appeared in the 5-7th and 15-20th days, being 14%/g and 20%/g. But, around 10th, the uptake rate decreased somewhat. The maximum uptake rate was 20%/g in 20th days. The uptake rates of some metal ions were with the somtimes keeping pace differentiation of embryonic cells. But sometimes not and division. The tendencies of the uptake rates of Se-75, Rb-83, Sr-85, Co-56 and V-48 were similar, indicating that the metal ions were accumulated in the callus during somatic embryogenesis. The uptake rate of Mn-52 was not only similar to other elements, but the variation was like normal distribution.     

利用mRNA差别显示技术分析枸杞体细胞胚发生早期基因的差别表达

本研究选用枸杞体细胞胚发生体系中的继代愈伤组织(对照)、胚性愈伤组织和早期胚体为实验材料,提取细胞总RNA,在12种锚定真核生物mRNA3'末端的OligodT12VN中,随机选用OligodT12GA为引物合成了以上三种材料的cDNA第一链,以此cDNA为模板,用随机引物进行PCR扩增,选择差别表达的片段。我们选用了OPA、OPH、OPK和OPB四组的60个随机引物对所得的c DNA进行了PCR扩增,得到了三个在体细胞胚发生早期组织中基因特异表达的片段。结果表明,在体细胞胚发生早期有胚胎发生特异性基因的表达,而且这种特异表达的基因在继代愈伤组织中没有表达,说明植物的体细胞胚发生过程就是细胞内基因差别表达的结果。 Abstract:Embryogenic calli and early embryo can be obtained from both auxin and auxin-free medium.The analysis of differential gene expression in early somatic embryogenesis has been hindered by above-mentioned material.The modifications of the recently described mRNA differential display method were reported and differential gene expression in early slmatic embryogenesis was analyzed.We have obtained three differential bands of cDNA in early somatic embryogenesis.The results indicate that gene expression has temperal and spalil order in early somatic embryogenesis of Lycium barbarum L.Plant somatic embryogenesis is the results of differential gene expression in cell.     

枸杞髓组织培养中体细胞胚胎发生与过氧化物酶同工酶分析

枸杞髓组织在 MS+6 - BA0 .1mg/ L+NAA0 .5mg/ L培养基上诱导愈伤组织发生。在 MS+6 - BA0 .1mg/ L+NAA0 .5mg/ L+CH50 0 mg/ L培养基上继代培养 ,再转入 MS+6 - BA2 mg/L +NAA 0 .5mg/ L的分化培养基上进行分化培养。显微观察表明 ,在培养过程中愈伤组织细胞由非胚性细胞转变为胚性细胞 ,直至发育成体细胞胚胎和完整植株 ;电泳结果显示 ,体细胞胚胎发生的各阶段 ,其过氧化物酶同工酶发生相应的变化。     

Intergeneric Somatic Hybrid Plants of Nicotiana tabacum L. and Lycium barbarum L. by Protoplast Electrofusion

Mesophyll protoplasts of Nicotiana tabacum L. and protoplasts from cell suspension of Lycium barbarum L. were heterofused by electrofusion with a frequency of ca. 4%--5%. One hundred cell lines were selected at random identified by isozyme analysis with peroxidase and superoxide dismatase, and the differences from their parent were found. Results indicated that 9 cell lines expressed enzymatic bands characteristic of both parents. Five of the 9 cell lines were highly morphogenic and regenerated numerous young shoots that manifested morphological traits specific to both parents. However, these shoots never grew up or regenerate roots. Esterase analysis of leaf material from the regenerants of 5 morphogenic hybrid cell lines demonstrated that two of them (NL4 and NL8) expressed an unique hybrid band which were not shown in either parents. Cytological observation on parental and NL4 hybrid cell lines revealed that the somatic chromosome number of NL4 varied from 58 to 80, significantly higher than that of either parents. Ribosomal DNA analysis of NL4 and NL8 showed that NL8 covered all fragments of both parents: NL4 did not have the fragments characteristic of Lyciurn barbarurn L. Both hybrids had new fragments, suggestive of intermolecular recombination of rDNAs of the parents. Four normal plants morphologically similar to tabacco parent were obtained from hybrid cell hne NL4, which survived after being transferred to soil. Cytological analysis of root-tips from one of the plants indicated that it has ca. 58 chromosomes. This paper also discussed the problems on the production frequency and incompatibility of somatic cell hybrid.     

枸杞多糖提取工艺的研究

采用热水浸提法从枸杞干果中提取枸杞多糖,经正交实验确定浸提时间、温度、料水比最优值分别为：5h、100℃、1：40;微波预处理超过25min时,多糖得率达15．91％;醇沉时当醇沉时间为4h、乙醇加入量为4倍时有利于枸杞多糖提取。     

枸杞生物技术研究进展

综述了国内外枸杞生物技术研究进展,包括了枸杞无性植株的再生、胚培养在育种中的应用;单倍体和多倍体,单细胞和原生质体培养技术;体细胞突变体筛选以及遗传转化再生植株的研究进展。     

鱼腥草体细胞胚胎发生和植株再生

目的:利用鱼腥草的叶片和叶柄为材料,进行体细胞胚胎诱导及植株再生研究。方法:运用正交设计试验,考察在改良的MS固体培养基上添加不同种类、不同浓度的植物生长物质组合及其配比对鱼腥草愈伤组织诱导、体细胞胚胎发生及植株再生的影响。结果:鱼腥草无菌苗叶片在含有2,4-D 1.0 mg/L 6-BA 0.5 mg/L的改良MS培养基上能诱导出胚性愈伤组织;胚性愈伤组织在含有6-BA 1.0 mg/L的改良的MS培养基上诱导体细胞胚的发生;叶柄在含有6-BA 1.0 mg/L改良MS培养基上直接产生体细胞胚。体细胞胚在改良的MS NAA0.1 mg/L 6-BA 1.0 mg/L的培养基上能够快速繁殖,形成大量不定芽,在不加任何激素的MS培养基上就可以萌发出不定根,发育为成完整植株,在MS IBA 1.0 mg/L的固体培养基上能够形成大量的根。结论:建立了鱼腥草体细胞胚胎发生及植株再生的体系。