不同类型水稻材料成熟胚组织培养力的比较

【目的】研制适应性广的不同类型水稻成熟胚再生技术。【方法】采用生产上应用价值高的不同类型水稻品种成熟胚为外植体材料,比较研究9个粳稻、9个籼稻和11个新近培育的超级杂交稻(两系或三系)或优势组合在不同培养基中的出愈、分化和再生等组织培养力表现。【结果】以M8为基本培养基,在添加30g·L-1蔗糖、8g·L-1琼脂和2mg·L-12,4-D(2,4-苯氧乙酸)相同成分的基础上,供试粳稻材料单独添加较低浓度的细胞分裂素,即0.3mg·L-16-BA(6-苄基嘌呤)可获得较高的出愈率,出愈率在89.2%-61.7%之间;而供试籼稻材料则需在添加2mg·L-16-BA和0.5mg·L-1KT(激动素)两种细胞分裂素的基础上,再添加1mg·L-1NAA(萘乙酸)生长素类激素,可明显提高出愈率,出愈率在100%-9.1%之间;而11个偏籼型超级杂交稻则表现在添加1mg·L-16-BA和1mg·L-1KT(激动素)两种细胞分裂素,即可获得较高的出愈率,出愈率在86.3%-40%之间。在此基础上,在添加了0.5mg·L-12,4-D和1mg·L-16-BA的MS分化培养基(粳稻)以及0.2mg·L-12,4-D和0.5mg·L-16-BA(籼稻和超级杂交稻)的分化培养基上,可分别获得59.5%-9.2%(粳稻)、87.5%-3.6%(籼稻)和43.2%-17.2%(杂交稻)的植株再生率。【结论】初步建立了适合相同水稻类型成熟胚高频再生技术。     

The Comparison in Tissue Culture Ability of Mature Embryo in Different Cultivars of Rice

In order to study the regeneration technology of mature embryos in different rice varieties,nine japonica,nine indica and eleven hybrid rice varieties of two line or three line or superiority combinations were selected as explants to study the callus induction,differentiation and regeneration rates on different media.The higher callus induction （61.7-89.2%） was observed in japonica rice,when cytokinin was added at lower concentration （0.3 mg L-1 6-BA） in M8 basal medium,supplemented with 30 g L-1 sucrose,8 g L-1 agar and 2 mg L-1 2,4-D.Further,the addition of two cytokinins （2 mg L-1 6-BA,0.5 mg L-1 KT） and 1 mg L-1 NAA in the M8 basal supplemented medium resulted in 9.1-100% of the callus induction in indica rice.The percent callus induction in hybrid rice varieties was 40-86.3% when addition of 1 mg L-1 6-BA and 1 mg L-1 KT was added,and the cytokinins was required by the japonica and indica rice varieties in the M8 basal supplemented medium.It was observed that when the 0.5 mg L-1 2,4-D and 1 mg L-1 6-BA were added in japonica rice,and 0.2 mg L-1 2,4-D and 0.5 mg L-1 6-BA were added in indica and hybrid rice in the MS different media,the regeneration rates were 9.2-59.5%,3.6-87.5% and 17.2-43.2% for japonica,indica and hybrid rice,respectively.Thus,the regeneration technology with higher output is established in the mature embryos of similar rice varieties.     

几个水稻粳型亲籼系成熟胚离体培养研究初报

对4个粳型亲籼系成熟胚离体培养特性研究的结果表明,种子贮存150d后的愈伤组织平均诱导率为82.02%,显著高于种子贮存20d后的诱导率(51.04%)。愈伤组织诱导率、愈伤组织质地结构、以及绿苗再生率在材料间表现出差异,其中材料间愈伤组织平均诱导率从高到低的顺序为G3004-4>G2605>G2417-2-1>G2416-3,材料之间绿苗再生率的顺序为G3004-4>G2416-3>G2605>G2417-2-1,表明供试的粳型亲籼系在S-b座位携带中性基因Sn比Si基因易于提高愈伤组织诱导率、改善愈伤组织质地结构,但S-b座位的基因型对绿苗再生率的影响不明显。不同培养基对各材料愈伤组织诱导率的影响不同,表现为在诱导培养基上添加1g·L-1水解乳蛋白和0.5g·L-1脯氨酸均能够显著提高G2417-2-1和G3004-4的诱导率;分化培养基上不同激素配比也影响其绿苗再生率。4个粳型亲籼系成熟胚离体培养特性类似于粳稻的培养特性。     

Dicamba and Sugar Effects on Callus Induction and Plant Regeneration from Mature Embryo Culture of Wheat

To establish a highly efficient plant regeneration system for wheat genetic transformation, the effects of three different concentrations of dicamba and two different sugar types on callus induction and plant regeneration from mature embryo cultures were evaluated. Callus induction and plant regeneration were obtained from mature embryos of two commercial cultivars Zhoumai 18 and Yumai 34 （Triticum aestivum L.） cultured on L3 basal medium. The results showed that the efficiency of mature embryo culture was significantly influenced by the genotypes, sugar types and dicamba concentrations. 4 mg L^-1 dicamba proved the best effective for inducing embryogenic callus and also gave the highest proportion of plants regenerated across the two cultivars. Substitution of maltose by sucrose significantly improved the plant regeneration efficiency in both cultivars. There was a significant interaction between genotype-by-sugar types, and sugar types-bydicamba concentrations. Overall, Zhoumai 18 gave the highest frequency of plant regeneration （82.65%） when dicamba concentration was 4.0 mg L^-1 and with sucrose in initial callus induction. These results will facilitate genetic transformation work with elite wheat.     

橄榄成熟胚培养研究

以橄榄常见品种惠圆的成熟胚为外植体，研究橄榄成熟胚培养的方法、培养反应和影响因素。建立了橄榄成熟胚离体无菌培养的有效方法；获得了橄榄无菌胚苗、愈伤组织、气生根和胚状体等4种培养物，为进一步的橄榄离体快速繁殖、细胞悬浮培养、原生质体培养、遗传转化以及离体种质保存等生物技术研究奠定基础。     

宁夏水稻成熟胚离体再生体系的建立

以34个宁夏粳稻品种的成熟胚为材料,N6和MS为基本培养基,研究了基因型、激素、分化,继代对宁夏粳稻成熟胚培养特性的影响.结果表明,基因型对宁夏粳稻成熟胚培养的影响较大,参试材料中优育28的培养特性最好;诱导愈伤组织中2,4-D质量浓度以2 mg/L较好;分化培养中参试品种均表现为KT 2mg/L+NAA 1 mg/L+IAA 0.5 mg/L处理的分化效果最好;成熟胚诱导的愈伤组织继代1～2次对分化没有显著的影响,继代3次以上,其分化能力明显减弱.通过本研究,初步建立了宁夏水稻成熟胚高频再生体系.     

甜玉米成熟胚和幼胚再生性比较分析

采用21种不同基因型的甜玉米,以成熟胚和幼胚两种不同外植体,对其愈伤组织的诱导、继代培养和分化进行了研究.结果表明,玉米的成熟胚和幼胚与愈伤组织诱导率相关,但两种外植体诱导出的愈伤组织状态明显不同,幼胚诱导出的愈伤组织状态普遍比成熟胚的好.对愈伤组织状态进行了分类,研究了出愈率、克隆力、芽点数与植株再生的关系,结果表明出愈率、克隆力与再生能力有较大的相关性.     

CuSO_4和烯效唑对寒地粳稻成熟胚培养的影响

为了建立寒地粳稻成熟胚的高频再生体系,以寒地粳稻空育131和东农425为试验材料进行成熟胚培养,研究CuSO4和烯效唑的单独和组合使用对水稻成熟胚培养的影响。结果表明:在培养基中单独添加15.0μmol/L的Cu2+或添加0.50 mg/L的烯效唑有助于寒地粳稻成熟胚的诱导继代和分化;在诱导和分化培养基中同时添加CuSO4和烯效唑时,空育131在CuSO4浓度为20.0μmol/L且烯效唑浓度为1.00 mg/L时,愈伤组织的诱导率和分化率都达到最高;CuSO4浓度为10.0μmol/L,烯效唑浓度为1.00 mg/L时,东农425愈伤组织的诱导率最高,CuSO4浓度为15.0μmol/L,烯效唑浓度为1.00 mg/L时,东农425愈伤组织的分化率最高。     

水培条件下硫素营养对粳稻米质的影响

 【目的】探讨水培条件下,硫素营养对粳稻稻米品质的改善作用,明确其品质指标随硫素浓度改变而变化的规律,以期提高人们对硫肥的重视,为硫肥的精确合理施用奠定理论基础。【方法】以粳稻武粳15和武育粳3号为材料,采用水培法,研究硫素营养对粳稻稻米品质指标的影响。【结果】在适宜营养液硫浓度下,武粳15和武育粳3号的稻米品质得到显著改善。武粳15和武育粳3号分别在200 μmol?L-1和260 μmol?L-1硫浓度处理下,加工品质、外观品质指标得到显著改善,与对照相比,胶稠度分别增加了12.1 mm和8.4 mm。在米粉糊化特性上,武粳15在200 μmol?L-1硫处理的米粉峰值粘度较对照增加143 cP,消减值降低290 cP。武育粳3号在260 μmol?L-1硫浓度处理下,米粉的峰值粘度较对照增长158 cP,消减值降低34 cP。两个品种籽粒蛋白质含量及总游离氨基酸含量随硫浓度的升高呈增加的趋势,4种蛋白组分含量与硫浓度总体上呈正相关关系。【结论】200 μmol?L-1硫处理的武粳15和260 μmol?L-1硫浓度处理的武育粳3号,其品质相关理化指标得到较好的改善。     

酸枣成熟胚离体培养研究

为了建立酸枣成熟胚离体培养再生体系,以酸枣成熟胚为试验材料,研究启动培养基和增殖培养基对不同贮藏处理酸枣成熟胚萌芽率及增殖的影响。结果表明：以常温保存1a的成熟胚萌芽率较高（57.78％）,最佳启动培养基为不添加任何植物激素的M S培养基,而最适增殖培养基为1/2MS＋1.0 mg/L 6 BA＋0.3 mg/L NAA＋1.5 g/L PVP。     

水分胁迫对粳型巨胚稻幼苗性状的影响

 为探寻提高粳型巨胚稻幼苗素质的方法,用PEG 6000配制成0,5%,10%,15%,20%,25%等6种不同质量浓度的溶液,对粳型巨胚稻西巨胚1号种子进行水分胁迫,用粳稻合系22-2（小胚）作对照,研究部分幼苗素质性状,试验方法不同,其结果差异较大：清水引发PEG液培养的水分胁迫处理不利于幼苗生长,随着PEG浓度的升高,幼苗生长势变弱,叶绿素和可溶性蛋白含量增加,根系活力下降,SOD活性在轻度、中度胁迫下上升,严重水分胁迫下呈下降趋势; PEG引发清水培养的水分胁迫处理在PEG质量浓度15%以下时对幼苗的生长有一定的促进作用,可溶性蛋白降低,SOD活性和根系活力升高,浓度高于15%后对幼苗生长、SOD活性和根系活力有抑制作用。西巨胚1号经5%～15%的PEG溶液引发24h后再用清水培养的水分胁迫处理下幼苗素质有所提高。     

水稻成熟胚不同接种方式对愈伤组织诱导的影响

采用4种不同的接种方式,对3个水稻品种的成熟胚进行了愈伤组织诱导试验。结果表明：在胚上纵切1刀,将种子接种在培养基上的接种方式,愈伤组织诱导率最高、出芽率最低,且愈伤组织质量最好,可以在短时间内获得大量胚性愈伤组织,建立悬浮系,其愈伤组织在较短的时间即可分化出再生植株,是1种简单、方便、效果好的诱导水稻愈伤组织的理想方法。     

小麦成熟胚的组织培养

【研究目的】为了筛选出小麦(TriticumaestivumL.)成熟胚(MEs)培养和遗传转化受体材料的优良基因型,【方法】以27份优良普通小麦冬性主栽品种或高代育种品系为试材,采用整粒切胚诱愈方法(endosperm-supportedmethod),对小麦成熟胚进行组织培养研究。【结果】结果表明,在成熟胚培养过程中光照、温度、培养基2,4-D浓度、基因型等因素对小麦成熟胚组织培养影响较大;同时小麦成熟胚组织培养存在着较强的基因型依赖性,不同基因型愈伤组织诱导率、分化率、成苗率的差异具有统计学意义(P<0.01)。通过研究从27个小麦基因型中筛选出了3个适合于成熟胚整粒切胚诱愈的基因型,即山农2618、泰山021、河北341,他们的成苗率分别达到了15.38%、25.27%、21.33%。【结论】小麦成熟胚组织培养优良基因型的筛选,为以后小麦的遗传转化和应用基因工程技术进行遗传改良奠定了基础。     

Study on Plant Regeneration of Wheat Mature Embryos Under Endosperm-Supported Culture

To reveal the suitability of using mature embryos as an explant source in wheat tissue culture, mature embryos from eight common wheat cultivars （Triticum aestivum L. cv.） were cultured with or without endosperm to test their efficiency of callus induction and plant regeneration. When embryos were cultured together with endosperm （endosperm-supported culture, ES）, the percentage of callus induction was significantly lower than that when embryos were cultured in the absence of endosperm （non-endosperm-supported culture, NES）. This pattern was evident in most genotypes, regardless of whether 2 or 8 mg L^-1 2,4-D was added in the NES culture. However, in ES culture, more induced calli were differentiated into distinct green spots and they further developed into plantlets. Thus, more plants were regenerated in ES culture than in the NES treatment. Most of the eight tested genotypes showed a significant difference in callus induction rate and plantlet regeneration in both ES and NES cultures. In addition, the enzymatic activity of oxalate oxidase in the callus of ES culture condition was obviously higher than that in the callus of NES culture condition, suggesting that the activity of oxalate oxidase may be a parameter for selection of calli with potential for plantlet regeneration. These results indicate that wheat mature embryos are valuable explants for highly efficient callus induction and plant regeneration, if proper treatment and medium are used.     

结缕草成熟胚愈伤组织的诱导及再生体系的研究

以结缕草的成熟胚为外植体,通过胚性愈伤组织诱导进行植株再生。结果表明在MS+2,4-D(2.0mg/L)+VB1(4mg/L)的培养基中,成熟胚可较高频率的诱导产生愈伤组织,诱导率为43.20%,但愈伤组织质量较差。将其在降低2,4-D浓度和添加6-BA,ABA的继代培养基中继代培养,可明显改善愈伤组织质量,增加胚性愈伤。筛选继代的胚性愈伤组织置于分化培养基MS+6-BA(1.0mg/L)+KT(2.0mg/L)+NAA(0.1mg/L)中,分化率45.6%。本研究建立了一套有效的以成熟胚为外植体结缕草组织培养再生体系。     

文冠果成熟胚离体培养及细胞学研究

以文冠果成熟胚为外植体,进行了初代培养、继代培养和生根培养研究,结果表明:初代培养诱导文冠果成熟胚形成不定芽的适宜培养基是MS+2.0mg.L-16-BA+0.1mg.L-1NAA+30g.L-1蔗糖+6g.L-1琼脂,分化率可达94.7%;继代高生长的适宜培养基是MS+0.5mg.L-16-BA+0.5mg.L-1NAA+30g.L-1蔗糖+6g.L-1琼脂,培养50d后,苗高大于0.5cm芽数量平均达11.67株;生根培养的适宜培养基是WPM+1.0mg.L-1IBA+30g.L-1蔗糖+6g.L-1琼脂,生根率为70.30%,平均根数为3.57条。并通过石蜡切片技术,观察愈伤组织诱导、愈伤组织分化不定芽以及不定芽发育过程。     

Establishment of a Highly Efficient Regeneration System for the Mature Embryo Culture of Wheat

Establishment of a highly efficient regeneration system for the mature embryo of wheat will provide a convenient tool for wheat tissue culture and transformation,thereby facilitating the transformation of foreign genes into wheat.By using the mature embryos derived from 20 different wheat lines including Shi 4185,Yumai 66,Lunxuan 987,CB037,Yangmai 6,Xinchun 9,Bobwhite,Han 6172,Zheng 9023,Jimai 20,Ningchun 4,and Jing 411,the effects of some factors including inoculation methods,initiating culture media,organic additives,antioxidants,and auxins on the regeneration from the explants were evaluated.The results indicated that the scraping embryo culture was better than the whole embryo culture,the Aa medium was better than the SD2 medium and dicamba was better than 2,4-D in increasing the regeneration frequency.An Adi medium was established in this study by adding silver nitrate,cysteine,ascorbic acid,dicamba,glutamine into the Aa medium at the concentration of 4,40,100,2,and 5 mg L-1,respectively.By using the Adi medium and the scraping technique,the regeneration frequencies of the mature embryos of CB037,Lunxuan 987,Han 6172,Yangmai 6,Bobwhite,Zheng 9023,Shi 4185,and Jimai 20 became 85.6,60.1,46.0,42.1,42.0,34.0,33.0,and 32.0%,respectively,which were about 5-8 times higher than that obtained from the conventional culture mediums and techniques.This novel regeneration system could be helpful in wheat transformation.     

高留胚率水稻辽粳294成熟胚组织培养条件的优化

以高留胚率水稻辽粳294成熟胚为外植体,对影响高留胚率水稻愈伤组织诱导、分化的接种方式、培养基中激素水平进行研究。结果表明,用解剖刀在胚上纵切1刀后再将种子接在培养基上,种子灭菌彻底,愈伤组织诱导率高且质量较好。诱导培养基中激素6-BA和KT的浓度对愈伤组织的诱导率有极显著的影响,诱导培养基为N6+2 mg/L 2,4-D+0.3 mg/L 6-BA+0.1 mg/L KT时,诱导率较高,可达90.0%;分化培养基为MS+0.8 mg/L NAA+2 mg/L 6-BA时,绿苗分化效果好,分化率为73.0%。     

白羊草成熟种子组织培养影响因素的研究

以白羊草成熟种子为外植体,研究不同表面消毒方法、外界条件及继代对愈伤组织诱导的差异。结果表明:外植体选用酒精浸泡30 s、0.1%升汞处理15 min为理想消毒方法;MSB+2.0 mg/L 2,4-D结合完全黑暗培养,愈伤诱导率达到86.7%;经继代培养基MSB+1.0mg/L 2,4-D继代后,分化率明显提高。     

小麦成熟胚离体培养研究

以美国小麦品种Overley成熟胚作为外植体进行离体培养,研究外源激素对小麦成熟胚诱导愈伤组织、分化出苗以及试管苗增殖的影响。试验结果表明,在MS培养基中添加2,4-D3mg·L-1愈伤组织诱导频率较高,成愈率为84.0%;在添加有KT2～3mg·L-1的MS分化培养基中愈伤组织分化率较高,可达85%以上;将试管苗转接至添加0.2mg·L-1的NAA和0.5～2.0mg·L-1的6-BA的培养基中,试管苗有一定的分化增殖,但最高分化率仅为33.3%,增殖系数2.05。