水稻离体育性变异研究

对野败型不育系珍汕９７Ａ、保持系珍汕９７Ｂ、恢复系ＩＲ２４、ＩＲ２６、泰引１号、明恢６３、红莲型不育系红源Ａ、包台型不育系包源Ａ、光敏核不育系农垦５８ｓ、温敏核不育系Ｗ６１５４ｓ等１０个水稻材料的幼穗在不同的培养基上培养、再生植株及对其后代进行育性鉴定,探讨了体细胞无性系变异中雄性不育突变发生的机率以及影响离体筛选雄性不育变异体因素。因素表明：在５个材料（珍汕９７Ｂ、红源Ａ、包源Ａ、Ｗ６１５４ｓ和     

光（温）敏核雄性不育水稻育性转换中幼穗和花药的某些生理指标变化

光（温）敏核雄性不育水稻（Ｐｈｏｔｏｐｅｒｉｏｄ／ｔｅｍｐｅｒａｔｕｒｅ－ｓｅｎｓｉｔｉｖｅｇｅｎｉｃｍａｌｅ－ｓｔｅｒｉｌｅｒｉｃｅ,简称Ｐ／ＴＧＭＲ）８９０２５和培矮６４ｓ在秋季从不育阶段经半不育阶段向可育阶段转变进程中,对其花粉母细胞形成期（Ⅴ）、花粉母细胞减数分裂期（Ⅵ）的幼穗和成熟花药（Ⅷ）中过氧化物酶（Ｐｅｒｏｘｉｄｓｅ,简称ＰＯＤ）、超氧物歧化酶（Ｓｕｐｅｒｏｘｉｄｅｄｉｓｍｕｔａｓｅ,简称ＳＯＤ）．和过氧化氢酶（Ｃａｔａｌａｓｅ,简称ＣＡＴ）活性测定表明,幼穗和花药相同发育时期的酶活性随正常花粉的比率升高而增加,但以成熟花药中增加最多．并伴有丙二醛（Ｍａｌｏｎａｌｄｅｈｙｄｅ,简称ＭＤＡ）含量减少．表明自由基代谢的变化可能与光（温）敏核雄性不育水稻花粉败育有关,且自由基代谢变化在幼穗发育中期即已开始．     

水稻体细胞无性系R_1、R_2代中的雄性育性变异观察

通过水稻幼穗培养,１９９１－１９９２两年间,在５个品种（珍汕９７Ｂ、红源Ａ、包源Ａ、Ｗ６１５４ｓ,和南广占）中共获得了５０株雄性不育变异株,其中Ｒ_１代有４８株,Ｒ_２代有２株。在Ｒ１代,共获得５２６８株再生植株,雄性不育变异的平均频率为０．９１％（０．８３－１．０８％）;在Ｒ_２代（珍汕９７Ｂ）发生雄性不育变异的频率为２％。本文报道了多种花粉败育类型之间可以相互转变现象,此外不育和可育之间亦可以相互转变。对离体培养产生的雄性不育变异株用一批现有ＣＭＳ（Ｃｙｔｏｐｌａｓｍｉｃｍａｌｅｓｔｅｒｉｌｅ）不育系的典型保持系、恢复系进行测交,结果表明,Ｗ６１５４ｓ产生的雄性育性变异株仍保持核不育的特性;红源Ａ产生的雄性育性变异株有的可能是嵌合体,有的其败育花粉类型虽发生了变化,但其恢保关系并没有改变,有的则可能已转成类似ＷＡ型的不育材料;南广占产生的典败变异株,其恢保关系类似ＷＡ型,可能属核不育转成ＣＭＳ的首例发现。     

水稻单,二倍体幼穗培养及其后代变异品系

水稻草倍体幼穗在含2,4－D的MS培养基上培养可以产生二倍体植株,并易再生成植株;经几代繁殖筛选,育出4个生物特性和农艺性状优于亲本的突变品系,田间试验表明,可用于农业生产上。     

甲基磺酸乙酯(EMS)对水稻离体幼穗影响的初步研究

用不同浓度的EMS处理水稻离体幼穗,测定培养初期的呼吸强度、细胞色素氧化酶活性和同工酶以及苹果酸脱氢酶同工酶,发现低浓度的EMS对细胞呼吸强度以及细胞色素氧化酶活性有刺激作用,而高浓度的EMS则表现为抑制效应。呼吸强度和酶活性与幼穗培养力之间具有平行关系。研究结果表明,EMS的原初生理效应是影响幼穗细胞的代谢过程,幼穗体细胞诱导频率的改变是幼穗培养初期供能系统受到影响的结果。     

小麦×冰草属间杂种F_1的植株再生及其变异

在对0.5—40.cm长幼穗培养4周后诱导出愈伤组织的基础上,获得了88株普通小麦(Triticum aestivum cv.Chinese Spring,2n=6x=42,AABBDD)×沙生冰草(Agropyron desertorum Fisch.> Schult.,2n=4x=28,PPPP)杂种F_1的再生植株。不同长度的幼穗在培养时,其愈伤组织发生的部位及其增殖速度不同。再生植株的产生主要是通过直接器官发生途径。所有的再生植株染色体数目全与杂种F_1相同,为2n=35。与杂种F_1相比,再生植株的减数分裂行为是相当复杂的,证明有染色体结构变异的发生。每茎穗数、叶片失绿斑等形态上的变异是由环境效应引起的;而株高、每穗轴节上小穗数和育性是由遗传效应决定的。特别值得注意的是,再生植株的自交结实率高达5.49%,共获得自交种子484粒,这对利用P染色体组中的期望基因有着极为重要的意义。     

影响水稻幼穗培养体细胞胚胎发生因素的研究

用水稻幼穗为外植体进行组织培养,研究了影响其体细胞胚胎发生的有关因素,建立了高频率体细胞胚胎发生的培养程序。结果表明不同基因型之间体细胞胚胎发生率的差异达到61．2％;生长素2,4－D对诱导水稻体细胞胚起重要的调控作用,细胞分裂素BA有一定的协同促进作用。干燥处理可提高愈伤组织体细胞胚胎发生率,愈伤组织含水量在60％～80％范围的培养效果较好;外源DNA溶液浸泡发育早期的心状体细胞胚．其进一步发育时异常胚的频率显著增加．成苗率降低。     

小麦幼胚培养再生单性雌花的研究

通过45个基因型的小麦（Triticum aestivumL.)幼胚培养,发现有11.1%的基因型从靠近再生芽基部的愈伤组织上分化形成花器官,再生花芽呈裸露的,多子房丛生的,具有藏盛羽毛状柱头而缺乏雄蕊,外稃,内稃和颖片的单性雄花。组织切片观察发现,其雌蕊起源于再生芽附近的分生组织细胞,并通过次生雌蕊再生的方式形成从生状,其羽毛状结构的发育先于子房中胚珠的分化,除正常的单胚珠外,还发现双生胚珠分化。X^2独立性检验结果显示,花芽再生率存在强烈的基因型效应,小麦品种YA-1表现突出（44.4%)。其花芽再生潜力能在不同年份间较好地再现,说明YA-1的花芽再生能力具有相对稳定性,与脱分化培养基的效应相比,YA-1的花芽再生效率主要受继代培养基成分的影响,其中,6-BA,NAA和加倍无机铁盐的配比较2,4,-D和正常浓度无机铁盐的配比更有利于YA-1的幼胚培养再生花芽,同时,外植体实验表明,YA-1的幼穗和成熟胚培养无任何成花反应,而其幼胚外植体具有特异的花芽再生能力。据此认为,YA-1的幼胚培养有助于为小麦花发育机理研究建立理想的实验系统。     

Variation in Anther Culture Response and Fertility of Backcrossed Hybrids between Indica and Japonica Rice (Oryza sativa)

This study was conducted to investigate the variations of fertility, callus induction and plant regeneration in backcrossed hybrids between a responsive japonica variety (Mankeumbyeo, recurrent parent) and a recalcitrant indica variety (Ranta Emas, donor parent) to anther culture. The mean spikelet fertility of inter-subspecific F₁ and BC₁F₁ hybrids was 33.5% and 67.2%, and the spikelet fertility of BC₁F₁ among backcrossed hybrids showed the most extensive variation (a low of 4.5% to a high of 90.6%). The mean fertility and distribution range in BC₅F₁ hybrids were almost the same as that of the japonica recurrent parent (94.6%). The mean callus induction of F₁ and BC₁F₁ hybrids was higher than that of donor parent, and the distribution range in BC₁F₁ hybrids varied from a low 0% to a high 18.7%. The mean callus induction and plant regeneration of BC₄F₁ hybrids was almost that of japonica recurrent parent, and there were no statistical differences between BC₄F₁ and BC₅F₁ hybrids. These results may help to accelerate the introgression of desirable traits from indica into japonica rice using anther culture of backcross hybrids as a breeding strategy.     

水稻花药培养植株后代的DNA变异

对籼稻圭６３０和粳稻０２４２８及其Ｆ１通过花药培养获得的８１个ＤＨ系进行了ＲＦＬＰ分析,有２８个探针揭示了ＤＮＡ变异。８１个ＤＨ系不同程度地发生了变异,并具有以下特点：（１）ＤＮＡ变异类型包括限制性片段长度的变化、９ＮＡ片段的丢失以及ＤＮＡ序列的扩增;（２）变异发生在籼稻圭６３０供体片段中的频率高于粳稻０２４２８,表现出基因型差异;（３）染色体组中第３、８、９和１０染色体较少发生变异,在其它染色体上均存在易变异位点;（４）在染色体的一些区段,相邻的探针均揭示了ＤＮＡ变异,表明在染色体上存在ＤＮＡ易变异区域;（５）变异位点和变异类型具有特异性,在同一位点不同的ＤＨ系中发生相同的变异。     

Studies on Cell Suspension Culture and Plant Regeneration in Rice

The present paper reports the establishment of rice cell suspension culture system, including callus induction and proliferation, isolation of single cells and small aggregates, cell suspension culture and callus re-formation, as well as regeneration of plantlets. The results have been obtained as follows: 1. The compositions of the different media used for callus induction, callus proliferation, cell suspension and plant regeneration are summarized in Table 1.2. Two kinds of disifectants, mercuric chloride and sodium hypochlorite, were used for surface sterilization of brown rice. The percentage of callus formation and callus yields were much higher when sodium hypochlorite was used (Fig. 3). We suggest that the disinfactant is one of the important factors that affect callus formed at the initial stage has an influence upon subsequent isolation of cells and suspension culture and even plant regeneration. 3. Table 3 shows that addition of yeast extract to the medium improves callus yield greatly and the efficiency of callus formation to a lesser extent. 4. Both medium Ⅱ (modified B5 medium) and N6 medium were suitable for cell suspension culture, but medium II was more effective for cell growth and callus re-formation (Fig. 4 and Table 4). 5. Effect of 2, 4-D on cell growth was tested at the concentration range among 0, 10-6, 10-5, 10-4 to 10-3 M. The results indicated that 10-5 M of 2,4-D was most effective for induction of rice callus. It has also been found that absence of 2,4-D increased callus re-formation in suspension culture, but no plant regeneration was observed. 6. By using 7% sucrose in differentiation medium, for all the three varieties, the plant regeneration frequency was raised up to 3 or 4 times than those of the 3% ones (Table 6). Occurrence of albino plants is often reported as one of the problems in rice anther culture. It is, however, no problem in seed-derived rice cell culture.     

The Preliminary Studies on Culture of Unfertilized Ovary of Rice in Vitro

The present paper reports the results of the culture of unfertilized ovaries of rice in vitro. The inducting medium was N6 supplemented with 2 mg/L 2,4-D, 500 mg/L casien hydrolysate and sucrose was 4%. The differentiated medium was N6 supplemented with 2 mg/L Kinetin, 500 mg/L casein hydrolysate and the concentration of the sucrose was 3%. The 4 cultivars and 2 crossed combinations were used as the experimental materials. The experiments were shown the differentiation of the callus occurred amony various cultivars. The induced frequency in the crossed combinations was higher than that in the cultivars. Now 12 green plants and 3 albino plantlets have been obtained. The chromasomes of 11 green plantlets have been examined. Among them, 6 plantlets were haploid (n =12 ) and 5 plantlets were diploid. The embryoids were located in the micropylar end. Some of them possessed the suspensor, similar as zygote embryos. The callus was found from different origin. One of them was originated from haploid tissue derived from the nuclear in the embryo sac. Another was originated from the diploid tissue in the integument or ovary wall. The origin of the callus from the unfertilized ovary was discussed.     

籼稻体细胞培养再生植株染色体变异的研究

以IR36及IR54等品种的成熟种子及幼穗为外植体,获得籼稻体细胞培养再分化植株,并研究了再生植株当代(即第一代,SC_1)的染色体变异。在319株SC_1植株中发现四倍体10株,占总数的3.1％。在二倍体中发现不育株7株(占2.2％),其中经细胞学分析发现2株(1984及1985年各发现1株)为多染色体相互易位杂合子。减数分裂的研究表明,MRT植株终变期时染色体构形呈十分复杂的情况。除正常的12 Ⅱ外,还呈现出一系列的多价体。配对最高价性为拾价体,7 Ⅱ+1Ⅹ的构形占各种染色体构形总数的50.7％,分布最多。在这类染色体构形中,拾价染色体或呈环形(以7 Ⅱ+1⑩表示),或呈链形(以7 Ⅱ+1(?)表示)。这表明该植株12对染色体中有5条非同源染色体发生了相互易位,而这两株植株正是这种染色体易位的杂合子。     

起源于组织培养的籼稻雄性不育体细胞无性系变异

IR8及IR54的成熟种子与幼穗培养在MS培养基,蔗糖6％附加生长刺激素及其它成分的条件下,在再分化植株第一代(R_1)中各发现了1株雄性不育突变(其中来自IR54的1株为育性嵌合体)。在IR24及IR54体细胞无性系第二代(R_2)各发现1个株系分离出雄性不育与育性正常两类植株。上述雄性不育有花粉败育与无花粉型两类。IR24对无花粉型的雄性不育(来自IR54)为半恢复。F_1及R_2代的育性分离完全符合15/16∶1/16的模式,表明雄性不育为两对独立的核基因所控制。通常雄性不育是通过杂交(包括远缘与品种间)理化诱变或自发突变产生的。最近在烟草上通过原生质体融合也获得了雄性不育。但从体细胞培养的无性系及其后代中获得雄性不育的突变,本研究乃属首次。     

起源于籼稻体细胞培养的联会消失变异的细胞遗传学研究

对表现不育的体细胞培养再生植株作减数分裂细胞遗传学分析发现,一株由IR54幼穗外植体起源的不育株(二倍体)为部分联会消失变异。其减数分裂早前期染色体配对正常,在终变期及中期Ⅰ观察到了数目不等的单价染色体,后期Ⅰ出现各种数目的落后染色体。由于减数分裂时染色体不平衡而导致该再生植株不育。