褪黑素对‘无核白’葡萄体细胞胚的诱导作用

以欧洲葡萄‘无核白’（Vitis vinifera‘Thompson Seedless’）未开放的小花蕾为外植体,研究0、1.0、2.0、3.0 mg ? L-1褪黑素（Melatonin）对其体细胞胚的诱导效果。结果表明：‘无核白’小花蕾在 MS + 1.0 ~ 3.0 mg ? L-1 褪黑素 + 2.0 mg ? L-1 6-BA + 30 g ? L-1蔗糖 + 3 g ? L-1植物凝胶的培养基上继代培养30 d时愈伤组织诱导率较好,为73.67% ~ 89.10%,显著高于2,4-D处理,愈伤组织形成所用时间较2,4-D处理缩短了14 d。愈伤组织诱导120 d时,不同浓度褪黑素均出现体细胞胚,其中,以MS + 1.0 mg ? L-1褪黑素 + 2.0 mg ? L-1 6-BA + 30 g ? L-1蔗糖的培养基体细胞胚的发生率最高,180 d时达12.05%。体胚在 MS + 60 g ? L-1 蔗糖 + 0.5 g ? L-1活性炭的X6培养基中萌发30 d后,将萌发的子叶胚转移至MS + 0.2 mg ? L-1 6-BA + 0.1 mg ? L-1 NOA + 30 g ? L-1 蔗糖 + 0.5 g ? L-1活性炭的成苗培养基上,其中,1.0 mg ? L-1褪黑素诱导产生的体胚发育正常的数量较多,为14.84%。不同浓度褪黑素处理的体胚诱导率均于180 d后增长较快,且低浓度的褪黑素有利于体胚萌发与正常发育,2,4-D诱导的愈伤组织无体胚形成。     

红姜花体细胞胚胎发生及植株再生的研究

以红姜花（Hedychium coccineum）的花丝和花药为外植体,通过体细胞胚胎发生途径建立了植株再生体系。结果表明：外植体在MS + 4 mg ? L-1 2,4-D + 4 mg ? L-1 NAA + 1 mg ? L-1 6-BA + 30 g ? L-1 蔗糖 + 7 g ? L-1 琼脂的培养基上经过120 d诱导出愈伤组织,愈伤组织在MS + 1 mg ? L-1 2,4-D + 0.25 mg ? L-1 NAA + 0.25 mg ? L-1 6-BA + 30 g ? L-1 蔗糖 + 7 g ? L-1琼脂的培养基上经过继代筛选,获得浅黄色松散易碎的胚性愈伤组织。胚性愈伤组织在MS无机盐 + B5维生素+ 100 mg ? L-1谷氨酰胺 + 230 mg ? L-1 脯氨酸 + 100 mg ? L-1麦芽提取物 + 0.02 mg ? L-1 NAA + 0.02 mg ? L-1 TDZ + 0.5 ~ 1.0 mg ? L-1 2,4-D + 45 g ? L-1蔗糖的培养基中通过3个月的悬浮培养,可得到均质稳定的胚性细胞悬浮系（ECS）。胚性悬浮细胞在SH无机盐 + B5维生素 + 100 mg ? L-1谷氨酰胺 + 230 mg ? L-1脯氨酸 + 100 mg ? L-1麦芽提取物 + 0.25 mg ? L-1 NAA + 0 ~ 0.20 mg ? L-1 TDZ + 45 g ? L-1蔗糖 + 7 g ? L-1琼脂的体胚诱导培养基中培养10 d,可见到白色半透明体胚发生,20 d后体胚发育成熟。当TDZ浓度为0.15 mg ? L-1时,体胚诱导率高达4 500个 ? mL-1 PCV ECS（PCV：细胞密实体积）。在SH无机盐 + B5维生素 + 0.20 mg ? L-1 IAA + 0.25 ~ 1.0 mg ? L-1 6-BA + 30 g ? L-1蔗糖 + 7 g ? L-1琼脂的体胚萌发培养基上,体胚萌发率高达100%。将萌发的体胚转移到1/2MS + 1 g ? L-1活性炭成苗培养基中,进一步发育成正常的再生植株,植株室外栽培成活率达90%。     

龙芽楤木体细胞胚状体发生及成苗的调控研究

利用龙芽楤木叶片、叶柄、嫩茎及根为试材,研究了2,4-D、6-BA、NAA、糖种类对胚性愈伤组织及胚状体的诱导、芽分化和成苗的影响。以2,4-D1.0 mg/L,确定诱导胚性愈伤组织及胚状体的最适外植体为嫩茎,诱导的胚性愈伤组织及胚状体质量、数量均高于叶片、叶柄和根;经直观分析、方差分析和多重比较,结合对比试验,结果表明:6-BA是龙芽楤木芽分化及成苗的主要因素。以蔗糖30 g/L作碳源,芽分化最适培养基为MS+6-BA1.0 mg/L+NAA0.02 mg/L;成苗最适培养基为MS+6-BA1.0 mg/L+NAA0.04 mg/L,芽分化率和成苗率达88.02%和79.62%。     

芹菜胚性愈伤的诱导及高频植株再生体系的建立

通过对芹菜离体培养过程中激素配比、基因型、不定芽产生等条件的研究,建立了芹菜高频植株再生体系。研究表明,芹菜苗期的下胚轴最适于诱导愈伤组织,其愈伤组织诱导和继代培养的最佳培养基是MS+2,4-D1.0mg·L-1+KT1.0mg·L-1和MS+2,4-D0.5mg·L-1+BA0.2mg·L-1+CH500mg·L-1+4%甘露醇;基因型对愈伤组织和胚状体的诱导影响很大;60%以上的愈伤组织在继代培养基上可以从非胚性状态向半胚性和胚性状态转化。愈伤组织在1/2MS+1.5%蔗糖+CH500mg·L-1+KT0.25mg·L-1上获得不定芽或小试管苗;在无激素的MS或1/2MS固体培养基上继续培养不定芽或小试管苗,即可生根或长大,获得芹菜的完整再生植株。     

枣幼胚愈伤组织诱导和胚状体发生

以陇东马牙枣盛花后50～60d的幼胚为试材,采用三步培养法诱导胚状体。结果表明:诱导愈伤组织适宜的培养基为Nitsch培养基上附加BA 1.0mg/L+2,4-D 1.5mg/L+蔗糖30g/L+琼脂4.0g/L。诱导胚状体培养基宜选用,MS+BA 1.0mg/L+IBA 0.2mg/L+LH0.5mg/L+VC 0.1mg/L+蔗糖30g/L+琼脂4.0g/L,并获得完整植株。     

东北百里香组培再生体系的建立

 以中国特有地被植物东北百里香为试材,研究了植物生长调节剂组合对腋芽萌发和茎段、叶片外植体愈伤诱导和不定芽分化的影响。结果表明：百里香茎段腋芽可直接诱导萌发,在MS + 6-BA 0.5 mg · L-1 + NAA 0.1 mg · L-1培养基上萌发率最高,达74%,在MS + 6-BA 0.5 mg · L-1 + NAA 0.1 mg · L-1的培养基上增殖倍数为36.43。由茎段萌发的组培苗在1/2MS + IBA 0.5 mg · L-1的培养基中20 d后生根率100%,移栽成活率76.7%。继代培养中叶片外植体可以诱导出愈伤组织,但是不能进一步分化成苗。茎段愈伤诱导的最适培养基为MS + 6-BA 0.5 mg · L-1 + 2,4-D 1.0 mg · L-1,再分化培养基为MS + 6-BA（0.1 ~ 1.0）mg · L-1 + GA3（0.1 ~ 0.5）mg · L-1,分化率为33.3%。     

中国特有濒危植物八角莲胚状体的发生及其植株再生研究

以我国特有濒危药用植物八角莲(Dysosma versipellis (Hance) M.Cheng)为试材,采用DPS软件正交设计法和SPSS的Duncan's multiple range test,研究了不同植物激素对其愈伤组织形成、胚状体诱导及植株再生的影响,以期为离体培养条件下快速诱导八角莲愈伤组织、胚状体及其植株的再生,以及将来深入研究真菌诱导子诱导八角莲活性成分鬼臼毒素累积的作用机理、信号调控机制和人工种子制作等提供参考依据.结果 表明:八角莲叶和叶柄最适合作为诱导愈伤组织的材料;植物激素对愈伤组织形成的影响效果从大到小依次为2,4-D＞ TDZ＞KT＞NAA＞2-ip,愈伤组织形成的最佳培养基为MS+2,4-D 1 mg·L-1 +NAA 0.05 mg·L-1 +TDZ 0.5 mg·L-1 +2-ip 1 mg·L-1;诱导颗粒状愈伤组织胚状体形成的最佳培养基为MS+6-BA 0.5 mg·L-1 +NAA 0.1 mg·L-1,诱导率为71.33％;胚状体生根和植株再生培养基为MS+IBA 0.5 mg·L-1 +GA30.5 mg·L-1.     

科技文摘

正南瓜未受精胚珠的离体培养及植株再生以南瓜(Cucurbita moschata)品种试验1号为材料,以未受精胚珠为外植体,研究了激素种类、外植体发育时期、高温预处理时间和AgNO3浓度对胚状体诱导的影响。结果表明,2,4-D、NAA和6-BA组合有利于胚状体的形成,出胚效果最好的培养基为MS+1.0 mg·L-1 2,4-D+ 0.25 mg·L-1 NAA+0.5 mg·L-1 6-BA,出胚率达31.1%;雌花开放当天的胚珠出胚率最高(26.7%),且愈伤组织形成频率低(5%);外植体在黑暗、高温(35℃)条件下处理5天有利于胚状体的形成,出胚率为32.2%。     

黄瓜(Cucumis sativusL.)花药培养

以卡罗尔、HH1-8-57与津绿农家乐3个不同基因型黄瓜品种为试材,在Kumar等、Song等和詹艳等试验研究基础上,设计试验分别探讨低温预处理时间、培养基成分(胚性愈伤组织诱导培养基、胚状体诱导培养基)和基因型等因素对黄瓜花药培养的影响。结果表明:在4℃低温预处理2d时,胚性愈伤组织诱导率显著高于其他处理;在MS+1.0mg·L-12,4-D+0.5mg·L-16-BA+3%蔗糖+0.8%琼脂培养基上胚性愈伤组织诱导率最高,达到81.3%;在MS+0.1mg·L-1NAA+3.0mg·L-16-BA+3%蔗糖+0.8%琼脂培养基上胚状体诱导率最高,为40.0%;基因型间胚性愈伤组织诱导率及胚状体诱导率差异显著,津绿农家乐品种胚性愈伤组织诱导率最高,达到81.1%,HH1-8-57胚状体诱导率最高,为40.0%。本研究从2份试材中成功地诱导出胚状体并获得了黄瓜单倍体再生植株。     

桂花胚的离体培养

 以不同发育时期的桂花胚为试材, 研究了不同生长调节剂组合、基本培养基、光照时间、蔗糖浓度对离体胚萌发, 以及不同培养基对萌发幼苗生根的影响。结果表明: 诱导胚萌发较理想的培养基为B5 + 6-BA 2.0 mg·L^-1 +NAA 0.1 mg·L^-1 +蔗糖3% ～5% , 光照12 h·d^{- 1} ; 生根较合适的培养基为B5 + IBA 2.0 mg·L ^{- 1} +蔗糖3%。     

酿酒葡萄‘神索’体胚发生及再生体系遗传稳定性分析

 以酿酒葡萄‘神索’未成熟合子胚为外植体, 通过植物生长调节剂、光照、温度等因素的控制, 研究了葡萄体胚的产生、保存及植株再生。结果表明, 以NN为基本培养基, 诱导胚性愈伤组织的适宜植物生长调节剂水平是1.0 mg·L ^{- 1} 2,4-D; 诱导体胚的生长调节剂水平是1.0 mg·L ^-1 NAA + 0.5 mg·L ^{- 1} BA, 体胚诱导率为37.5%; 5℃微光条件, 适宜体胚的保存; 体胚成熟及植株再生的植物生长调节剂水平是0.05 mg·L ^{- 1} NAA + 0.5 mg·L ^{- 1} BA, 成苗率为42.1%。利用流式细胞仪并结合染色体计数对体胚及再生植株细胞核DNA含量及染色体鉴定表明, 体胚细胞染色体存在一定的变异, 而体胚再生植株倍性稳定, 细胞核DNA含量及染色体数与供体母株一致。     

‘月月红’月季体细胞胚胎发生和植株再生研究

 以建立月季高频再生体系为目标,以‘月月红’月季叶片为外植体,探索了生长调节剂和光照条件对其体细胞胚胎发生及其植株再生的影响。结果表明：3.0 mg · L^-12,4-D + 0.5mg · L^-1 TDZ的胚分化培养基上暗培养12周后转入胚成熟培养基,置于红光（光强约为7.2 µmol · m^-2 · s^-1）条件下培养,能产生较多具肥厚子叶且有高频再生潜力的体细胞胚。暗培养条件下主要产生无子叶的芽状胚。 胚性愈伤组织在含     

Somatic embryogenesis and plant regeneration in Psidium guajava L. cv. Banarasi local

A protocol for plant regeneration by somatic embryogenesis was developed in guava cv. Banarasi local by using immature zygotic embryo explants. Best induction of somatic embryogenesis was achieved from 10-week-old zygotic embryos on MS medium supplemented with 2,4-d (4.52 μM) and 5% sucrose. Maximum number of somatic embryos was produced when zygotic embryo explants were transferred to growth regulator free full strength MS basal medium after 8 days treatment with 2,4-d. Full strength MS basal medium containing 5% sucrose was most favorable for maturation of somatic embryos. Highest frequency of conversion and normal plantlet production were recorded from elongated torpedo stages of somatic embryos on half strength MS medium containing 3% sucrose. Over 90% of rooted shoots survived acclimatization.     

仙客来体细胞胚发生和发育过程中淀粉粒的变化

 以仙客来（Cyclamen persicum Mill.）开花植株的新生叶片为外植体，诱导筛选胚性愈伤组织，并使其进一步发育成体细胞胚。组织切片观察结果表明：胚性愈伤组织由胚性和非胚性细胞组成，胚性细胞多以胚性细胞团的形式存在，胚性细胞团起源于诱导的胚性决定细胞。体细胞胚起源于单个胚性细胞，经多细胞原胚、球形胚、心形胚和鱼雷胚等时期发育成完整植株。在体细胞胚的发生发育过程中，淀粉粒出现4次积累高峰，分别为胚性细胞、球形胚、早期鱼雷胚和成熟胚发育成完整的小植株时期，淀粉代谢与体细胞胚发生、发育及小植株的形态建成密切相关。     

梅花不同外植体离体培养及体细胞胚诱导植株再生

以梅花品种‘雪梅’和‘绿萼’的叶片、叶柄、茎段及未成熟子叶为外植体进行离体培养,探讨了体胚发生途径与植株高效再生方法。结果表明,成熟组织再生能力低,不定芽再生困难。在添加1.0mg·mL-1BA,1.0mg·mL-1NAA,1.0mg·mL-1IBA的1/2MS培养基中,‘雪梅’和‘绿萼’未成熟子叶的体胚诱导率最高,分别为40.4%和25.3%;同时将初生胚转移至新鲜培养基上可产生大量次生胚。在添加0.5mg·mL-1BA,0.1mg·mL-1TDZ和1.0mg·mL-1IBA的1/2MS基本培养基中,体细胞胚发育成完整植株的频率较高,‘绿萼’达26.7%,‘雪梅’达23.8%。再生植株成功移栽至温室且生长良好。     

西洋梨叶片直接再生体细胞胚

 西洋梨‘红安久’和‘绿安久’两个品种的无菌苗幼叶为试材, 研究了不同培养基对叶片分化不定梢及体细胞胚的影响。结果表明: 基本培养基的组成是影响叶片分化为不定梢还是体细胞胚的关键因素, 植物生长物质的种类和水平是影响体细胞胚再生率高低的重要因素。诱导培养基NN69 附加TDZ 2 mg/L 及NAA 0. 1 mg/L , 体细胞胚再生率最高, 绿安久达55. 5 % , 红安久为47. 3 %。NN69 附加BA5 mg/L 及NAA 0. 1 mg/L , 不定梢再生率最高, 绿安久达100 % , 红安久为85. 7 %。     

Cryopreservation of coriander (Coriandrum sativum L.) somatic embryos using sucrose preculture and air desiccation

An efficient protocol for cryopreservation of somatic embryos of Coriandrum sativum, an important spice and medicinal herb, was developed. The successful cryopreservation procedure utilized embryo clumps (ECs) comprised of 3–4 somatic embryos at the globular or heart-shape stage. These ECs were precultured for 3 days on medium supplemented with 100 g/L sucrose, desiccated under the current of sterile air for 100 min, then sealed in cryovials and plunged directly into liquid nitrogen. Preliminary incubation on sucrose-enriched medium (100 g/L) improved both desiccation- and cryo-tolerance of ECs compared to medium with normal sucrose content (30 g/L) and enhanced embryo formation after cryopreservation. The regrowth after cryopreservation and average number of new embryos developed from cryopreserved ECs were retained at the level of the untreated control (98% and 13 embryos per clump, respectively). Both normal and abnormal plants were produced from control and cryopreserved cultures, indicating that appearance of abnormalities was not related to cryopreservation. The regenerants with normal phenotype showed the same peaks of relative DNA content regardless of cryopreservation. The results suggest that simple desiccation method is effective for cryopreservation of coriander somatic embryos with subsequent regeneration.     

Regeneration of different Cyclamen species via somatic embryogenesis from callus,suspension cultures and protoplasts

The present study is the first report of the establishment of embryogenic callus cultures from seedling tissue, the regeneration of plants via somatic embryogenesis and the development of a regeneration system from protoplast to plant, using three wild species of Cyclamen, Cyclamen graecum Link, Cyclamen mirabile Hildebrand, Cyclamen trochopteranthum Schwarz (syn. Cyclamen alpinum hort. Dammann ex Sprenger). The ability to form embryogenic callus and to regenerate via somatic embryogenesis was strongly genotype-dependent for each species. From 0.5 g callus, up to 1461 somatic embryos were formed in the case of C. mirabile. Culture media with different concentrations of plant growth regulators, CaCl₂ and activated charcoal significantly influenced embryo formation in this species. Up to 1.4 × 10⁶ protoplasts were isolated from 1 g of C. graecum cell suspension. Diverse growth responses of the protoplasts in two embedding agents, agarose and alginate, were observed for the different Cyclamen species. These specific growth characteristics could be used as a selection marker for future fusion experiments. From both protoplast culture systems, somatic embryos were regenerated, grown to plantlets and acclimatised to greenhouse conditions.     

河套蜜瓜子叶诱导形成体细胞胚的研究

 以河套蜜瓜成熟种子子叶为外植体, 研究体细胞胚途径再生培养的主要影响因子, 并对体胚发生过程进行了观察。结果表明; 体细胞胚发生对培养基糖源浓度要求较高, MS附加60 g/L的糖源效果最好; 最佳植物生长调节剂组合为2,4-D 2.0 mg/L + 6-BA 1.0 mg/L; 1日苗龄子叶为最佳外植体源; 在诱导培养基上经暗培养1周、光培养1周, 再转入胚发育培养基, 有利于成熟胚的产生; 体细胞胚可产生在愈伤组织内部或表面、外植体表面、不定根上及外植体维管束的束中分生组织等部位。     

Somatic embryogenesis in a broad spectrum of grape genotypes

Somatic embryogenesis is the preferred method for cell-to-plant regeneration of grapevine. In this study, we tested the embryogenic capacity of anther-derived calli from 59 grape genotypes, representing a diverse group of Vitis vinifera and hybrid varieties, and hybrids and accessions of non-vinifera Vitis species. Most genotypes were tested on two types of media: MST1 medium, which contained plant growth regulators (PGRs) 2,4-dichlorophenoxyacetic acid (2,4-D) and thidiazuron (TDZ), and MSE medium, which contained 2,4-D and 6-benzylaminopurine (BAP). Twenty-four of the grape genotypes produced embryogenic callus on one or both of these media, eighteen of which have not been reported to form somatic embryos before. The results also suggested that the various PGR combinations are differentially effective at inducing somatic embryos in various classes of grape genotypes. For example, seven of the eight V. vinifera conv. occidentalis varieties brought forth somatic embryos on MSE medium, and three out of four American Vitis genotypes produced somatic embryos on MST1 medium. We could not observe any apparent association between frequency of callus formation and embryogenic capacity of the anthers.