小麦花粉管生长途径及受精过程经历时间的研究

应用常规石蜡切片和荧光显微镜技术对小麦花粉管生长途径及受精过程相应时间进行研究。结果表明，授粉后，花粉随即萌发，2个精子进入花粉管，营养核留在花粉粒内，不久解体。花粉管进入羽毛状柱头分支结构的细胞间隙，继续生长于花柱基部至子房顶部的引导组织的细胞间隙内。随后穿过子房壁，在子房壁与外珠被之间的缝隙中向珠孔方向生长。自花粉萌发至花粉管长入珠孔大约需要1 h。自花粉萌发并到达柱头分支结构中，花粉管均具明显的绿色荧光；但在引导组织区以及子房壁与外珠被之间生长的花粉管几乎看不到荧光。授粉后1 h花粉管经珠孔及珠心表皮细胞间隙进入1个助细胞，并释放精子。授粉后2～6 h精卵融合，授粉后16 h合子第一次分裂。授粉后2～3 h精子与极核融合，授粉后4 h初生胚乳核第一次分裂。授粉后2～16 h，即精卵融合至合子分裂前为花粉管通道法转化合适的时间，采用花柱横切滴加法转入外源DNA。     

Use of irradiated pollen as mentor pollen to induce self-fertilization of two self-incompatible Upper Amazon cacao clones

Summary Two self-incompatible Upper Amazon cacao clones, T85/799 and T79/501, were pollinated with compatible Amelonado pollen subjected to varying doses of gamma irradiation (10–100 Gy). The proportion of flat non-viable beans to fully formed, viable beans in the pods increased with an increase in dosage of gamma rays. At 60 Gy all the beans produced were flat and non-viable, beyond this dosage fruit set was zero. Pollinating the self-incompatible cacao clones with a 1 : 1 mixture of compatible mentor pollen irradiated at 60 Gy and normal self pollen produced a mixture of flat, non-viable beans and fully-formed viable beans. Similar experiments using irradiated pollen with a marker gene suggested that the fully-formed viable beans resulted from selfing. Increasing the proportion of the radiation-treated compatible pollen in the mixture increased the number of fully-formed beans. However, when compatible pollen which had been treated either at 80 Gy or with temperatures of 35° C, 40° C and 45° C for periods of five, ten and fifteen minutes in factorial combination were mixed with self pollen, no successful pollinations were achieved. Pollen viability tests indicated that, whilst pollen treated at 60 Gy were about 50% viable, those treated at either 80 Gy or with temperatures of 35–45° C were mostly not viable. This suggests that, to overcome the incompatibility in cacao, the tubes of the mentor pollen grains used should at least grow into the style. The possible causes for overcoming the self-incompatibility in cacao are discussed.     

A procedure suitable for in vitro pollen selection inBrassica oleracea

Summary Experiments to determine whetherBrassica oleracea pollen could effect fertilisation following incubation in liquid culture medium are reported. Pollen was incubated in vitro, collected by centrifugation and used to pollinate compatible pistils. While retaining viability and the capacity to germinate such pollen was unable to penetrate the stigma papillae. However, incubated pollen produced functional tubes following pollination of styles from which the stigmas had been removed. These tubes grew through the style to the ovary and viable seed was obtained. The potential application of this procedure in pollen selection is discussed.     

Viability and storage of bromeliad pollen

Several bromeliad species from two different subfamilies, were used to develop a reliable method to evaluate pollen viability. Pollen germination on a medium containing 20% sucrose, 0.001%H₃BO₃ and 0.5% agar was comparable to germination on a compatible stigma. Maximum germination was reached within 2 to 10 hours depending on the species. Based on this test, six species were considered as being good pollen donors with germination percentages between 49%and 83%. Furthermore, pollen from these species and cultivars could be stored in liquid nitrogen (–196 ^°C) without a considerable loss of viability. For all species, a dehydration period of 4 hours prior to cryopreservation and a rehydration period of 1 hour after cryostorage were essential. Greenhouse humidity influenced anther moisture content and cryostorability. This revised version was published online in July 2006 with corrections to the Cover Date.     

美国梾木(Cornus spp.)花粉活力及萌发特性

为了了解美国梾木花粉活力及萌发特性,采用扫描电镜技术对13个美国梾木无性系花粉形态进行观测与方差分析,并采用荧光显微技术和TTC染色法对无性系2号花粉萌发特性及花粉活力进行观察。结果显示:(1)不同无性系间花粉大小差异不显著,但外壁纹饰差异较大,极面沟距的变异系数最高且遗传最为稳定。(2)花序开花率为75%时花粉活力最高,整花4℃条件下短期保存花粉活力较高。(3)授粉后,多数花粉可在柱头上正常萌发,72 h后花粉管即可进入子房。研究认为,花粉外壁及极面沟距可作为鉴定美国梾木无性系的形态指标,美国梾木花粉活力及萌发特性是抗逆及丰产杂交组合的重要保证。     

Pollen tube growth and fertilization after different in vitro pollination techniques of Aechmea fasciata

The effect of cut style and placental pollination on fertilization efficiency was studied for compatible Aechmea fasciata plants. These alternative in vitro pollination techniques resulted in lower penetration rates of the ovules by pollen tubes in comparison to pollination on the stigma. An explanation was found in the intervention of the normal pollination process,through which less pollen-pistil interactions were built up. After cut style pollination the percentage of ovule penetration by pollen tubes increased when a longer style part was left at the ovary. Probably fewer factors that control pollen tube growth are present in the lower style part. Pollen germinated on the ovules after placental pollination but only rarely penetration of the micropyle by a pollen tube occurred. Activation of the ovary, induced by in vivo prepollination for 6 hours, and pollination two days after anthesis did not increase the fertilization percentage. Grafting a style with active growing pollen tubes to ovules on the placenta (placental grafted style pollination) resulted in a higher fertilization percentage. Pollen tube growth through the style was essential for pollen tube guidance to the ovules and penetration of the micropyle. This revised version was published online in August 2006 with corrections to the Cover Date.     

万寿菊花粉活力及柱头可授性研究

为探明造成万寿菊雄性不育系结实率低的原因,对万寿菊花粉活力及柱头可授性进行了研究。采用花粉离体萌发法研究万寿菊自交系V-01花粉萌发适宜温度、花粉活力日变化和适宜贮存条件;用联苯胺-过氧化氢法检测万寿菊雄性不育系S-261、S-17-06-29、S-0191不育株柱头的可授性;水溶性苯胺蓝染色法检测花粉在柱头的萌发情况。结果表明：（1）万寿菊V-01适宜萌发温度介于25~30℃之间;V-01花粉活力日变化趋势为先升高后降低,11-13时采集的花粉萌发率最高;4℃干燥贮存是最适宜的花粉贮存条件。（2）万寿菊柱头形态呈‘γ’状时有可授性,可授性可持续3天。（3）万寿菊花粉授粉到柱头上1 h内即可萌发,授粉后2 h,花粉细胞达到花粉管内。     

Storage of avocado pollen

Summary Avocado pollen was stored at a range of temperatures and relative humidities (RH) for up to one year and the pollen was tested for viability in vivo.Pollen stored for one month was capable of germination on the stigma and penetrating the ovule when stored at 4°C with <1,23,55 and 75% r.h. and at -196°C with 0% r.h. Most pollen samples stored at 25 and -15°C at a range of r.h. would germinate on the stigma but none would penetrate the ovule.After one year of storage, pollen at 4°C and <1 and 23% r.h. would germinate on the stigma but would not penetrate the ovule. There was no germination of pollen stored at 4°C and 55 and 75% r.h. Only pollen stored at -196°C and 0% r.h. would penetrate the ovule, but thawing and refreezing once during the year destroyed the viability.     

亚麻受精过程及其经历的时间

应用石蜡制片技术, 观察亚麻的受精过程及其各阶段经历的时间。结果表明, 亚麻开花时落到柱头上的花粉随即萌发, 花粉管在花柱引导组织的细胞间隙中生长, 进入子房后经子房内表面, 沿胎座经珠柄进入珠孔, 进入1个助细胞, 在卵细胞与中央细胞之间释放2个精子;精子脱掉细胞质鞘后,形成精核与细胞质体;2个精核同时分别进入卵细胞和中央细胞, 并与卵核及次生核融合;精核与次生核融合速度稍快;观察到合子中雌、雄性核仁融合的过程。受精各阶段经历的时间为,花粉落到柱头上随即萌发;开花后4.5 h左右, 花粉管长入胚珠的珠孔;4.5~5.5 h,花粉管进入1个助细胞并释放精子;5.5~6.5 h为精卵融合和精核与次生核的融合期;7.5~8.5 h初生胚乳核分裂;12.5 h之后,合子分裂;6.5~12.5 h为合子静止期。     

The breeding system of rubber (Hevea brasiliensis): an evaluation of controlled hand pollination methods

Summary Controlled hand pollinated pistils of rubber were observed using fluorescence microscopy to assess the efficiency of the universally-employed method for the production of progeny for plant breeding. The controlled hand pollination method conducted in the morning resulted in the deposition of a mean of 15.6 pollen grains on the stigma, with no stray pollination observed. Over 36% of the pistils had the potential to set fruit.Pollinations conducted in the afternoon at the normal time of anthesis, had double the fruit set potential of morning pollinations as measured by penetration of ovules by pollen tubes. Pollinator efficiency also varied, with excessive damage to stigmas resulting in reduced pollen germination and tube growth. There were differences between clones in both female and male fertility, in the proportion of pistils with more than three carpels and in the production of small abnormal stigmas. There was no difference in pollen tube growth following self- or cross-pollination, indicating that the self-sterility mechanism of rubber operates in the ovary. Pollen could be stored for 5 days at 5°C and 75% RH with a 22% loss of fertility.     

Storage of sugarbeet pollen

Summary To develop the technology for long-term pollen preservation, sugarbeet pollen was collected from plants grown in the greenhouse and in the field, and was stored 1 day to 1 year at 5, -18, and -196°C. Pollen containing about 12% moisture was successfully stored in liquid nitrogen (LN₂) up to 1 year; this pollen effected fertilization of male-sterile flowers as well as freshly collected pollen. Germination of the resultant seed was good and not different from seed from fresh pollinations. Pollen stored at -18°C for 1 year did not result in as much seed set as fresh pollen, and 1 year at 5°C was essentially lethal. In vitro pollen germination served as a post-storage viability measure, provided the pollen was hydrated before germination. The methods tested in these experiments provided a relatively simple, reliable, and inexpensive means for preservation of sugarbeet pollen for breeding purposes and for preservation of genetic resources.Joint contribution of the Agricultural Research Service, USDA, and the Beet Sugar Development Foundation.     

Effects of high temperatures on mature pollen grains in wild and cultivated maize accessions

Summary The effects of high temperature on mature pollen of various maize lines were investigated. Genotypic differences in pollen reaction to high temperature were revealed. Pollen grains resistant to high temperature (35°C, 26°C) were characterized by higher germination capacity and better ability to develop normal pollen tubes. The studies are of interest to evaluate reproductive system tolerance and conduct gamete selection at the mature pollen grain stage in maize.     

Hybrid Tea-rose pollen. I. Germination and storage

Summary Germination and storage trials were carried out with pollen of several rose varieties. The pollen grains germinated well in a 15% sucrose solution with 40 ppm boric acid. Staining the pollen with a 0.1% tetrazolium solution and standardizing the degree of colour at which the pollen grains are counted as viable, provided a good viability estimate, simpler to carry out than in vitro germination. Germination capacity and staining ability of the pollen were greatly impeded-about halved-by dehydration during storage in desiccators at low humidity. This effect could be corrected by humidifying the pollen beforehand for about one hour, though this pre-treatment increased the percentage of germinated pollen grains more than the percentage stained. There was no difference between the two percentages in fresh or in deep-frozen pollen.Pollen stored at 1°C and high relative humidity soon lost its germination capacity: between 0 and 20% humidity a considerable proportion of the pollen remained viable for 9 months and longer. Storage for the same period in vacuum-sealed glass tubes at –24°C maintained viability as well or better and would probably prolong it further. Some of the cold-stored pollen induced a reasonable seed set after one year, a low seed set was obtained even after two years of storage at 1°C and low humidity.     

Factors affecting haploid production in wheat using the Hordeum bulbosum system. 1. Genotypic and environmental effects on pollen grain germination,pollen tube growth and the frequency of fertilization

Summary A quantitative examination of pollen grain germination and pollen tube growth within the overy wall was made in selected wheat × tetraploid H. bulbosum hybridizations, to investigate the effect of the wheat and the H. bulbosum genotype on these characters. As expected, variation at the known crossability loci had no effect on pollen grain germination. The frequency of pollen tube penetration of the ovary wall was, however, severely reduced when the dominant alleles were present. Pollen tube penetration was nevertheless observed in 3 of the 110 ovaries examined of the non-crossable cultivar Highbury. The H. bulbosum genotype had a much smaller effect on these characters, but significant differences between the clones were observed in the numbers of pollen tubes initially penetrating the ovary wall. Although two H. bulbosum genotypes showed no significant differences in the number of ovaries with pollen tubes at the base of the ovule, significant differences in the frequency of fertilization were observed. The possible cause of this discrepancy is discussed.The frequency of fertilization in crossable wheat × H. bulbosum hybridizations was improved by the application of gibberellic acid within 10 minutes of pollination, and reduced by an increase in the ambient temperature from 20°C to 26°C. Fertilization following the pollination of non-crossable wheat genotypes was not affected by either of these factors.     

Pollen grain germination and pollen tube growth following in vivo and in vitro self and interspecific pollinations in annual Cicer species

Summary Germination of pollen grains and growth of pollen tubes were studied to determine the cause of barreness in crosses among annual Cicer species. In vivo and in vitro time-course studies and fluorescent microscopy revealed no pollination incompatibility among the selfs, crosses and reciprocals of C. arietinum L., C. reticulatum Lad. and C. cuneatum Rich. In general, Cicer pollen grains germinated and grew on styles of Cicer species. Pollen tube growth was characterized by irregularly spaced and intermittent callose deposits. Failure of seed formation in interspecific pollinations may be attributed to the slowness of pollen tube growth or collapse of fertilized ovules. In addition to these causes, shortness of stamens and sparsity of pollen grains were responsible for flower drop in natural selfs. Although the number of pollen tubes entering the micropyle in interspecific pollinations was low, it may be possible to grow the fertilized ovules on an artificial medium to obtain F₁ plants.     

Competition between 2x and x pollen in styles of Solanum tuberosum determined by a quick in vivo method

Summary Pollen competition in vivo was studied by placing two pollen samples on one stigma and comparing the length of the tubes under a U.V. microscope. Comparisons involving pollen of Solanum tuberosum and S. phureja showed that 2x pollen grew faster than x pollen, both in 2x and x styles. Consequences for induction of dihaploids in S. tuberosum are discussed.     

Pollen germination,pollen tube growth and fertilization following self and interspecific pollination of Paspalum species

Summary Crossability between most Paspalum species is very low. This study was undertaken to identify the impediments to hybridization. Accessions of P. intermedium Munro. ex Morong, P. jurgensii Hackel and P. dilatatum Poir were self-pollinated and crossed with one anther. Paspalum intermedium is essentially self-sterile but P. jurgensii and P. dilatatum are highly self-fertile. Following pollination, pollen germination and tube growth were studied by observing the pollinated pistils with fluorescent microscopy. Examination of self-pollinated pistils revealed that the pollen germinated shortly after contacting the stigmas. Germination was over 80% for the P. intermedium and P. dilatatum accessions but only 57% for P. jurgensii. Pollen tubes grew to the micropyle within 45 minutes after pollination in P. dilatatum and 1 hour and 15 minutes in P. jurgensii. However, in the P. intermedium accessions most tubes did not grow beyond the stigma and very few penetrated the style and ovary. Apparently stylar-incompatibility is the reason for the low selfed seed set. In the cross-pollinations, pollen germinated shortly after pollination and germination ranged from 57 to 88% for the different crosses. In all crosses the pollen tubes grew to the micropyle within 30 minutes to 2 hours after pollination indicating that a cross-incompatibility system is not the cause for low crossability among these species. By examining embryo sacs from P. intermedium × P. dilatatum, its reciprocal and P. intermedium × P. urvillei crosses, it was determined that gametes failed to unite in some crosses and this is a major reason for low crossability.     

Studies on the self-incompatibility of Petunia hybrida in excised-style culture. An attempt at improving a technique in excised-style culture

Summary A technique of excised-style culture was developed. In twelve compatible combinations of pollen and style, a correlation was found between the number of pollen tubes protruding from the style and the number of mature seeds formed in vivo. This suggests that the technique of improving excised-style culture is effective not only for studying the interactions between style and pollen tubes during tube growth, but also for predicting the production of seeds in vivo.     

麻疯树花粉管荧光染色及生长过程的研究

花粉管的生长是完成植物受精的重要生理过程,观察花粉管生长首先需要对雌蕊组织进行软化透明。本研究探索适合麻疯树雌蕊结构的透明方法,并利用苯胺蓝染色技术,在荧光显微镜下观察授粉后麻疯树花粉管的萌发和生长过程。结果表明:麻疯树雌蕊组织可采用NaOH溶液软化透明,较优的条件是2 mol·L-1的NaOH,50℃恒温箱内软化3~4 h;麻疯树授粉2 h后花粉管开始萌发,10 h后花粉管穿过花柱基部,11 h后在子房中观察到花粉管。麻疯树授粉后,花粉管进入子房至少需要11 h,完成受精则需要更长的时间。     

Compatibility and incompatibility in witloof chicory (Cichorium intybus L.). 4. Formation of self-seeds on a self-incompatible and a moderately self-compatible genotype after double and triple pollinations

Summary After double or triple pollinations including selfing and the use of one-day-old compatible foreign pollen a severely self-incompatible clone could be forced to develop a reasonable amount of self-seeds per flower head. This number of self-seeds increased when the foreign pollination was delayed for e.g. 240 minutes in the double pollination experiment. Even more self seeds were obtained in the triple pollination experiment with a delayed second selfing.The influence of the old foreign pollen may be caused by a mentor pollen or a pioneer pollen effect.