碳源组份及浓度对小麦花药培养和游离小孢子培养的影响

通过对小麦花药培养基中各种碳源的研究,发现单独使用麦芽糖作碳源的培养基出愈率最高,麦芽糖与蔗糖搭配使用培养效果次之,而单独使用蔗糖作碳源出愈率最低.在整个碳源比较试验中,随着培养基中麦芽糖比例的增加,出愈率随之提高.糖源对出愈率的影响与其水解产物有关.在游离小孢子培养中,麦芽糖是最好的糖源,其他糖源经培养未形成胚状体.检查培养3 d后的小孢子活力发现唯有麦芽糖作碳源的培养基有活力;认为高浓度的蔗糖葡萄糖以及葡萄糖与果糖混合糖源对小孢子有毒害作用.葡萄糖起始浓度达到2.0 mmol/L时,对小孢子有致死作用.当麦芽糖培养1 d后,再加入高浓度的葡萄糖发现小孢子存有活力,而且随着培养天数的增加,小孢子的活力也增加.     

单倍体育种技术在油菜育种材料创新上的应用研究Ⅱ.胚状体培育及试管苗越夏

利用游离小孢子培养技术对9个甘蓝型杂交油菜品种(组合)的F1代进行离体小孢子培养.在NLN-13培养基中诱导胚状体发生,将胚状体放置于含有多菌灵和链霉素的培养基中培养,结果表明:①20mg/L浓度的多菌灵和链霉素可以提高胚状体的成苗率.②将胚状体接种于含有0.05mg/L 2,4-D和0.01mg/L6-BA的MS培养基中培养,愈伤组织能快速分化形成不定根和丛芽.     

杂花苜蓿高效再生体系的建立

通过对新疆塔城杂花苜蓿品种下胚轴再生植株的系统研究，经筛选获得种子萌发培养基为：1／2MS无糖培养基；胚性愈伤组织诱导培养基：MS＋2，4-D2mg／L＋6-BA0．5mg／L＋蔗糖3％。胚性愈伤组织诱导率为100％；胚状体诱导培养基：SH＋2，4-DZing／L＋6-BA0．5mg／L＋CH250mg/L＋蔗糖1％，胚状体诱导率为86．96％；胚状体萌发培养基：SH＋GA30．5mg／L＋CH250mg／L＋蔗糖1％，胚状体萌发率为90％-95％；生根培养基：1／2MS＋IBA0．5rag／L＋蔗糖1％。生根率为100％。完成再生时间120d左右。     

小白菜小孢子胚状体诱导影响因素研究

以10个小白菜杂交种为试材,采用游离小孢子培养方法,研究影响小孢子胚状体形成因素的结果表明,不同品种胚状体的诱导率不同;NLN培养基中大量元素减半有利于小孢子胚诱导,其诱导率达到100%;培养基中添加0.05 mg/L的6-BA和0.2 mg/L的NAA可以提高胚状体诱导率;活性炭和琼脂糖对诱导胚状体发生和发育有促进作用,可使子叶胚的比例达90%以上;花蕾经4℃低温预处理24 h后的成胚率显著提高。     

红花玉兰胚性与非胚性愈伤组织的诱导及生理生化差异分析

为了得到生长状态良好的胚性愈伤组织(EC),本研究以红花玉兰的未成熟种子作为外植体,通过1/2 MS基本培养基添加不同浓度的2,4-D、6-BA、水解干酪素(CH)以及提高蔗糖浓度和使用脱落酸(ABA)两步法的措施来诱导胚性愈伤组织更好的生长,并通过测定其可溶性蛋白、可溶性糖以及游离脯氨酸等指标来进行差异分析。结果表明:最适合诱导胚性愈伤组织的培养基为1/2 MS+2.0 mg/L 2,4-D+0.2 mg/L 6-BA+750 mg/L CH(诱导率高达96.8%);蔗糖浓度提高到5%和6%时,胚性愈伤组织的状态较好,诱导率可达50.0%和64.7%,且测得胚性愈伤组织的可溶性蛋白和游离脯氨酸含量均显著高于其他处理,当蔗糖浓度为6%时,胚性愈伤组织的可溶性糖含量显著高于其他处理。结果表明胚性愈伤组织较非胚性愈伤(NEC)具有较高的代谢活性,且经高浓度蔗糖处理后的胚性愈伤对这一刺激作出了积极的响应,为体胚分化提供了科学的理论基础。     

火龙果花药培养诱导胚性愈伤组织及遗传稳定性

成年态火龙果茎段诱导的愈伤组织再分化较困难,进而影响到遗传转化效率。以红皮紫红肉的"紫红龙"火龙果花药为外植体,研究了小孢子不同发育时期、低温预处理时间、不同植物生长调节剂及浓度配比对花药诱导胚性愈伤组织的影响。结果表明,小孢子发育处于单核初期至单核靠边期的花药,其愈伤组织诱导率最高可达23.9%,但未能诱导出胚状体;而4℃低温预处理48 h,愈伤组织的诱导率可达32.8%;最适花药愈伤组织诱导的培养基为MS+2.0 mg/L 6-BA+1.0 mg/L 2,4-D+60 g/L蔗糖,诱导率为37.4%,胚性愈伤组织为黄绿色、致密状,外表有颗粒状突起。愈伤组织增殖培养基以MS+0.4 mg/L TDZ+0.8 mg/L KT效果较好,增殖系数达9.1倍以上。愈伤组织再分化培养基为MS+0.4 mg/L TDZ+0.5 mg/L 2,4-D+0.6 mg/L NAA时,愈伤组织再分化率达36.7%。通过火龙果花药诱导出愈伤组织,增殖获得大量胚性愈伤组织,再分化形成不定芽,经SRAP分子标记检测遗传性一致,能为火龙果基因工程育种和种苗快速繁育提供新的途径。     

基因枪转化光温敏雄性不育小麦受体的选择与培养条件优化

为了选择适宜的基因枪转化光温敏雄性不育小麦的受体,并对幼胚组织培养条件进行优化,以小麦光温敏雄性不育系BS210、BS366以及常规品种京411的幼胚为材料,分别对碳源、除草剂浓度和基因型进行筛选.结果表明,在以蔗糖和麦芽糖为碳源的诱导培养基上,幼胚的出愈率差异不显著;在以蔗糖和麦芽糖为碳源的分化培养基上,愈伤组织的分...     

苦瓜未授粉子房的胚状体诱导

苦瓜常规育种获得纯合株系耗时长、成本高,通过离体雌核培养途径获得单倍体可加速苦瓜育种进程。为建立苦瓜未授粉子房离体培养再生体系,本研究利用苦瓜未授粉子房为外植体,研究基因型、接种方式、激素配比、胚囊发育时期、预培养处理对胚状体诱导效果的影响。结果表明:MS+6-BA 0.5 mg/L+NAA1.00 mg/L+2,4-D 1.0 mg/L+TDZ 0.04 mg/L培养基胚状体的诱导率最高,为19.60%;在8个苦瓜品种中,2个植株生长势强的品种‘桂农科3号’和‘泰国山’苦瓜胚状体诱导率较高,适合作为离体雌核诱导的实验材料;开花当天的子房去除表皮后横切成2 mm的薄片可有效降低愈伤组织的形成,提高胚状体的诱导率;在黑暗和33℃条件下热激3 d有利于子房的转绿和出胚。本研究对苦瓜未授粉子房的胚状体诱导关键因子进行优化,为创制苦瓜单倍体育种材料提供帮助。     

小白菜小孢子胚状体诱导影响因素研究

以10个小白菜杂交种为试材,采用游离小孢子培养方法,研究影响小孢子胚状体形成因素的结果表明,不同品种胚状体的诱导率不同;NLN培养基中大量元素减半有利于小孢子胚诱导,其诱导率达到100%;培养基中添加0.05 mg/L的6-BA和0.2 mg/L的NAA可以提高胚状体诱导率;活性炭和琼脂糖对诱导胚状体发生和发育有促进作用,可使子叶胚的比例达90%以上;花蕾经4℃低温预处理24 h后的成胚率显著提高.     

提高冬小麦花药愈伤组织诱导效率的研究

为了进一步提高小麦花药愈伤组织的诱导效率,以12个黄淮麦区冬小麦杂交F1代为供试材料,比较了不同诱导培养基、培养方式和花药接种密度与共培养子房数等因素对花药愈伤组织诱导及分化的影响。结果表明:CHB培养基诱导效果最好,C17次之,而N6和W14效果较差,不同基因型对不同培养基的反应不同,但大多数材料在CHB和C17培养基上反应较好;培养基附加较低浓度的2,4-D(0.5 mg/L)时有利于形成胚状体并在诱导培养基上一步成苗;液体培养时愈伤组织诱导率最高但对愈伤的进一步分化不利,固体和双层培养时的诱导率最低,而半固体培养时不仅有利于愈伤组织诱导而且对其分化较好,是小麦花药培养的较好方式;花药与子房共培养可以显著提高愈伤组织诱导率,花药接种密度较低(2～2.4枚/mL)时较多的子房(20个)可以起到显著的促进作用,花药接种在高密度(6～7.2枚/mL)或中密度时(4～4.8枚/mL)10个或10个以上的子房诱导效果较好。     

The Effect of Carbohydrate Composition and Concentration on Anther Culture Response in Barley (Hordeum vulgare L.)

Culture medium composition is critical for the successful induction of microspore division in vitro. The present experiments have focused on a relatively neglected area of cell and tissue culture research, namely the carbohydrate component used in the medium. Three spring barley genotypes were cultured on a medium which was modified by replacing sucrose with the following carbohydrates (6 % w/v): maltose, fructose, malt extract, galactose and a glucose (3 % w/v)/fructose (3 % w/v) mixture. Both maltose and malt extract were superior to sucrose in their capacity to induce green plantlet differentiation from microspores. The concentrations of both sucrose and maltose were also varied. Overall the response of anthers on maltose based media was higher than on sucrose based media. Furthermore, a concentration of maltose m the range 6—12 % w/v produced a higher frequency of green plants than a low concentration (1—3 % w/v). The effect of maltose based media on germplasm of direct relevance to barley breeders was also tested. The cultivar ‘Blenheim’ was shown to be very responsive and this genetic factor was transmitted to the F₁ hybrid. The frequency of haploid to diploid regenerants was not consistent over genotypes, but in general there were more haploid than diploid regenerants. The implications of these results for barley breeding are discussed.     

Fertile maize lines obtained from isolated microspores

Microspore response of three- way cross maize hybrid genotype 3AL/95 (Zea mays L.) was studied under simplified isolation and culture conditions. Fertile plant production was achieved through abundant plant regeneration. As a total, microspores of 160 tassels were inoculated and five sustainable microspore derived callus cultures (SMC) were obtained. Hybrid seeds (ML SC), which were produced by crossing of regenerates from two SMCs, gave rise to subsequent vigorous and fertile progeny. The response of the 3AL/95 and ML SC microspores was studied in three liquid culture media in order to improve the early viability of microspores. Them N6M medium provided better survival of cultured microspores (p = 5%) than the ppN6M/89 and the YPM-G media. The pH 5.8 in mN6M medium revealed significant increase (p = 1%) in microspore viability as compared to pH 3.0. The ML SC microspores showed higher viability(30%) on the first day of culture in the mN₆M than those of the3AL/95 (19%) but without improved rate of callus formation and plant regeneration. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effect of Sugars in Wheat Anther Culture Media

Sugars are critical components in bread wheat (Triticum aestivum L.) anther culture media for successful somatic embryo initiation and plant regeneration. In this experiment, anthers from three wheat genotypes were cultured on a modified Liang's 85D12 initiation medium with seven sugar combinations (I-sugars: galactose, mannose, maltose, fructose, sucrose, glucose, maltose + glucose) at 0.26 M, and 2,4-dichlorophenoxyacetic acid (2 mg/L), 1-naphthalene acetic acid (1 mg/L), and glutamine (254 mg/L). Wheat starch (5 % W/V), a potential source of sugars, was used as the medium gelling agent. No previous research has studied the effect of different sugars with wheat starch. A split-plot experimental design with 42 replications was used with genotypes as whole plots and sugar combinations as subplots. Galactose and mannose did not support embryoid initiation and were dropped from the analysis. Averaged over the three genotypes, maltose was the best sugar (105 embryoids/100 anthers), followed by glucose (47 embryoids/100 anthers) and maltose + glucose (37 embryoids/100 anthers). These three sugar combinations were superior to the standard medium sugar, sucrose (24 embryoids/100 anthers), and to fructose (12 embryoids/100 anthers). The embryoids were divided into two groups for plant regeneration. The first group was transferred to regeneration medium (Liang 85D12 salts, sugars at 0.06 M, and wheat starch at 7 % w/v as gelling agent) with the same sugar (R-sugar) used as in initiation. The second group was transferred to regeneration media with sucrose. I+R-maltose (0.55)     

光温条件及碳源对苎麻愈伤生长和分化的影响

本试验用湘苎2号为材料,研究了不同光照时数和碳源、不同温度和碳源对苎麻愈伤生长和分化的影响,方差分析结果表明,用葡萄糖作碳源比蔗糖更有利于苎麻愈伤生长;12h光照/12h黑暗最有利于苎麻愈伤的生长,其次是全光照条件,黑暗条件下愈伤生长最慢。用葡萄糖作碳源时,愈伤生长量在30℃、28℃、25℃条件下均极显著大于23℃条件下;蔗糖作碳源时,在25～28℃的温度范围内有利于愈伤组织的增殖,而过高或者过低都是不利的。光照处理对分化是有利的,而相对较高温条件虽然能比较好的促进愈伤增殖,但这种来源的愈伤组织分化再生不如相对较低的温度条件下生长的愈伤。     

樱桃再生体系研究

摘要: 以樱桃枝条为材料,研究了樱桃离体培养及再生体系,探讨了不同激素组合对樱桃愈伤组织的产生、分化、增殖以及生根的影响,对樱桃再生体系进行优化,筛选出适合樱桃枝条腋芽诱导的培养基为MS+BA1.0mg/L+NAA0.1mg/L;无菌芽各种外植体形成愈伤组织、分化和增殖的培养基为MS+BA0.5mg/L+2-4D0.5mg/L;壮苗培养基为MS;生根的培养基为1/2MS+NAA1.0mg/     

国槐离体培养及高效再生体系的优化

以国槐枝条为材料,研究了国槐离体培养及再生体系,探讨了不同激素组合对国槐无菌芽的产生、愈伤组织的形成、分化、增殖以及生根的影响,筛选出适合国槐枝条无菌芽诱导、无菌芽各部分形成愈伤组织、分化和增殖的培养基为MS+ BA1.0mg/L +NAA 0.1mg/L;有利于生根的培养基为1/2MS+ NAA0.3mg/L。     

陆地棉原生质体培养与植株再生技术研究

 分离培养陆地棉品种ZDM-3（Gossypium hirsutum L cv ZDM-3）胚性细胞悬浮系的原生质体,通过体细胞胚胎发生成功获得再生植株。试验结果表明,植物生长调节剂组合和碳源对原生质体培养具有重要的影响,添加2,4-D + KT + 1.5%（W/V）葡萄糖+ 1.5%（W/V）麦芽糖的KM8P培养基对于陆地棉原生质体培养的效果较好。激素组合0.46 μmol·L^-1 KT + 0.45 μmol·L^-1  2,4-D诱导的愈伤组织较松软,分化潜力高;胚性愈伤组织的增殖使用0.93 μmol·L^-1 KT + 2.46 μmol·L^-1 IBA的激素组合;1.2 μmol·L^-1 IBA + 1.39 μmol·L^-1 KT有利于体细胞胚胎发生,而MSB-5 + 0.67 μmol·L^-1 KT+2.69 μmol·L^-1 NAA的培养基适合体细胞胚胎萌发和植株再生。此外,愈伤组织诱导宜用葡萄糖作为碳源,而胚性愈伤组织增殖、保存和胚状体的萌发过程宜使用麦芽糖作碳源。     

马蹄金(Dichondra repens Forst.)组织培养和快速繁殖

为了探究马蹄金组织培养及快繁技术,本研究以马蹄金为材料,使用75%的酒精和10%NaClO对不同外植体(胚根,下胚轴,子叶)进行消毒处理,在不同激素浓度配比的MS培养基中进行丛生芽诱导、愈伤组织诱导及分化,成功建立了马蹄金的组培快繁体系。结果表明:利用马蹄金下胚轴为外植体直接诱导不定芽效果较好,其最适程序为:流水冲洗30 min+75%酒精30 s+10%NaClO 9 min,污染率为5%,成活率为75%;下胚轴不定芽诱导的最适培养基为MS+1 mg/L 6-BA+0.3 mg/L NAA,诱导率为93.55%,出芽指数为8.86;胚根愈伤组织诱导的最适培养基为MS+1 mg/L 6-BA+0.5 mg/L NAA,诱导率为100%,下胚轴愈伤组织诱导的最适培养基为MS+0.5 mg/L 6-BA+1 mg/L NAA,诱导率为95.24%,子叶愈伤组织诱导的最适培养基为MS+0.3 mg/L 6-BA+0.5 mg/L NAA,诱导率为100%;愈伤组织不定芽分化的最适培养基为1/2MS+1.5 mg/L 6-BA+0.5 mg/L NAA,芽诱导率为20%,平均芽数为12.83;组培苗诱导生根的最适培养基为不加激素的1/2MS培养基;生根苗以瓶盖全开的炼苗方式移栽到以河沙为基质的育苗盒中成活率为100%,且添加蒸馏水与自来水对植株生长并无明显影响。本研究成功建立了马蹄金组织培养与快繁体系,为马蹄金种质资源改良及优质种苗生产提供了技术参考。     

色素万寿菊杂交母本未授粉子房培养

以色素万寿菊雄性不育株未授粉子房为外植体,比较研究子房发育期、高低温处理和生长调节剂组合对愈伤组织诱导和分化的影响。结果显示:发育6~10 d,花丝刚好露出花萼,顶部呈圆柱状的花蕾愈伤组织诱导效果最好,为79.2%;低温预处理3 d,高温培养5 d有利于愈伤组织诱导;MS+2,4-D 0.5 mg/L+NAA0.5 mg/L+6-BA 0.5 mg/L+KT 0.5 mg/L为愈伤组织诱导最佳培养基,诱导率高达85.8%;NAA 1.0 mg/L+6-BA3.0 mg/L为不定芽分化适宜培养基,分化率达77.6%;1/2MS+NAA 0.2 mg/L为生根适宜培养基,生根率93.5%。本研究可为建立高效单倍体培养体系以及单倍体育种技术提供基础性资料。     

Effect of genotype,environment and carbohydrate on anther culture response in head cabbage (Brassica oleracea L. convar. capitata (L.) Alef.)

Summary The effect of genotype, growing conditions for donor plants and type and concentration of carbohydrate in the culture medium was investigated for anther culture of head cabbage (white cabbage, savoy cabbage, pointed-headed cabbage). Strong genotypic effects on embryo formation from the cultured anthers were shown as well as superior embryo formation from anthers of field grown donor plants compared to plants grown in the greenhouse. When comparing 7, 10 and 13% sucrose in the medium, embryo response increased with increasing sucrose concentration. With maltose, which was generally inferior to sucrose as carbohydrate source for anther culture, the embryo response did not increase with maltose concentration above 10 per cent.