抗除草剂转基因水稻花粉漂移距离及生态风险分析

以抗除草剂转基因水稻Bar68-1及其杂种香125s/Bar68-1为花粉源,探讨了转基因水稻的花粉漂移距离和漂移频率,结果表明:在花粉源大(667 m2)、花粉受体为可育植株时转基因水稻的最大花粉漂移距离＜30 m,最大漂移频率0.295%;当花粉源小(4 m2)、花粉受体为部分可育的两系不育系时,最大花粉漂移距离＜9 m,最大漂移频率4.518%.因此,通常情况下,转基因水稻向正常可育的栽培稻转移基因的可能性小.另据分析,普通野生稻向栽培稻转移基因的可能性极低;湖南和江西栽培稻向普通野生稻转移基因的可能性小,两广和海南地区需要采取距离和障碍隔离防止栽培稻向普通野生稻转移基因.     

海南普通野生稻的花粉育性观察及杂交亲和性

对海南3个不同居群的普通野生稻(O.rufipogon Griff.)的花粉育性进行了观察研究,并采用荧光显微镜技术观察了转基因水稻花粉在不同居群的普通野生稻柱头上的萌发和在花柱内的生长过程。结果表明,以转基因水稻为对照,3个不同居群的野生稻相互之间花粉可育率在1%水平上差异极显著。用荧光显微镜观察时发现,转基因水稻花粉能在3个居群的普通野生稻的柱头上萌发并能到达胚珠,二者杂交亲和,同时发现,花粉管进入三亚居群胚珠的数目比儋州、万宁居群的数目多,说明相对于儋州、万宁居群来说,三亚居群与转基因水稻的杂交亲和性更高。     

抗除草剂转基因水稻基因漂移至常规栽培稻的频率研究初报

以抗除草剂草铵膦转基因bar水稻99-1作为花粉供体,四周种植常规栽培稻为花粉受体,观察抗性基因漂移频率。在距离转基因水稻2m以内,基因漂移频率为0.0193%～0.3960%,均在1%以下,但转bar基因抗除草剂水稻基因漂移至不同品种上的频率存在明显差异。     

抗草甘膦转EPSPS大豆的基因漂移研究

以抗草甘膦转EPSPS基因大豆和不同生长类型的非转基因大豆为材料,通过表型筛选及PCR分子检测,分析不同方向和距离条件下转基因大豆中外源基因通过花粉向非转基因大豆的漂移率,为制定转基因大豆安全种植隔离措施提供依据。结果表明,不同生长类型大豆的基因漂移率明显不同,以育成品系ZZH015最高,在70 612株中的漂移率为0.45%;半蔓生地方品种DPZ722次之,164 449株中漂移率为0.01%,而195 088株野生大豆YDD080中没有检测到基因漂移。方差分析表明,漂移率在不同距离和方向条件下均有显著差异,随着距离的增加不断递减。在5 m处漂移率为0.03%,而在29 m处降至0.001%。基因漂移在下风口方向发生较多,表明风媒可能是影响基因漂移分布的重要因素。如果以1%基因漂移率为阈值,无论是自然条件下还是随机选取1 000粒检测,漂移率都低于阈值,表明转EPSPS基因大豆的基因漂移可以忽略不计。     

甘蓝型油菜与诸葛菜杂交时花粉与柱头的识别反应

用扫描电镜观察了甘蓝型油菜与诸葛菜属间杂交过程中花粉粒与柱头间的认识反应，结果表明，正反交的花粉与柱头间认识反应是完全不同的．油菜柱头接受诸葛菜花粉后，个别提前萌发的花粉管能进入柱头，但当花粉大量萌发后，花粉管则不能进入柱头，在柱头表面出现爬行、扭曲、肿胀、产生分枝等不亲和现象．诸葛菜柱头接受油菜花粉后，多数花粉不能萌发，少数花粉管一旦萌发，即能顺利进入柱头。     

耐除草剂转基因大豆对花粉生活力的影响

转基因作物的环境安全评价是其商业化推广前的必要环节,其基因漂移风险及其可能引起的生态环境效应是安全性评价的重要内容。为了明确转EPSPS、PAT基因抗草甘膦大豆S4003. 12和S4003. 14外源基因的逃逸风险,于大豆盛花期选取温室种植的转基因大豆、其受体品种及常规栽培大豆材料花粉,使用液体培养基培养,检测其萌发率,并于带标尺显微镜下观察其花粉粒直径。结果表明:大豆花粉刚离体后萌发活力最强,培养2 h后花粉萌发率基本达到最大值约88. 2%,随离体时间延长萌发活力随之下降,室温放置3 h后,花粉几近完全失活;与非转基因对照相比,转EPSPS、PAT基因耐除草剂大豆S4003. 12和S4003. 14的花粉离体后的生存能力无显著差异,对花粉粒直径大小无显著差异。研究表明耐除草剂转基因大豆花粉传播能力与非转基因大豆间无显著差异,通过设置隔离区可有效规避外源基因逃逸风险。     

转基因花生外源基因田间漂移风险初步研究

用含花生条纹病毒壳蛋白(cp)基因和潮霉素(hygr)筛选基因的转基因花生品系为材料,田间试验表明转基因花生中的hygr基因可在短距离范围内随花粉漂移,离转基因花生种植区边缘1.0m 和3.0m范围内漂移频率分别为4.4%和5.0%;3.0 m以外范围内,没有检测到hygr基因随花粉漂移发生。各距离范围内均没有检测到cp基因随花粉漂移现象。因此认为,转基因花生外源基因漂移距离在3.0m以内。     

抗除草剂转基因水稻对稻田杂草种群的影响

草铵膦作为抗除草剂转基因水稻的专用除草剂,对千金子、节节菜、陌上菜、丁香蓼、鳢肠、双穗雀稗、四叶萍和水竹叶表现出优良防效,但不能有效防治牛筋草、牛毛毡、矮慈姑、野荸荠和空心莲子草。在田间使用草铵膦可以有效防治早期出苗的无芒稗,但不能防治中后期出苗的莎草科和阔叶类杂草。与常规稻秀水11相比,转基因水稻99 1的每穗实粒数显著高于秀水11,从而获得了更高产量。转基因水稻嘉禾201与亲本丙94 02比较,对稻田主要杂草密度和生物量的影响差异不显著。转基因水稻99 1和嘉禾201对无芒稗的影响因子R显著大于化感潜力品种地谷,但与非化感品种秀水11相当,表明99 1和嘉禾201无化感竞争优势。     

水稻丙酮酸脱羧酶基因OsPDC3功能的初步研究

 植物花粉中存在着活跃的有氧发酵过程。丙酮酸脱羧酶（PDC）作为发酵途径中的关键酶，参与花粉中的能量和物质代谢，在花粉萌发过程中起重要作用。通过反向遗传学的方法对水稻丙酮酸脱羧酶基因OsPDC3的功能进行了初步分析。OsPDC3是一个单外显子基因，编码蛋白与另外4个水稻PDC蛋白间的序列一致性达到77%~82%。表达模式分析显示OsPDC3在花粉中特异表达，GUS组织染色进一步证实其启动子具有花粉特异表达活性。超量表达OsPDC3会使转基因植株叶片中的PDC酶活性上升，表明OsPDC3在体内具有活性功能，能够参与花粉内的生理活动。反义抑制下调了OsPDC3在花粉中的表达水平，但对花粉的离体萌发率没有产生影响，推测这是由于PDC基因间的冗余造成的。     

抗虫耐除草剂转基因玉米花粉生活力研究

使用离体培养法检测不同玉米材料花粉萌发率并于显微镜下观察花粉粒直径,研究转Cry1Ab、EPSPS基因玉米对花粉生活力的影响。结果表明,刚离体花粉萌发活力最强,培养180 min后花粉萌发率可达89.4%,高温(36℃)条件下放置2 h花粉基本丧失萌发能力。与非转基因对照相比,转Cry1Ab、EPSPS基因玉米对花粉离体后的生存能力无显著影响,对花粉粒直径大小无影响。     

水稻雄性核不育突变体ms7的遗传分析及基因定位

【目的】通过对水稻雄性不育突变体的研究,可以鉴定更多与育性或花粉发育相关的基因,有助于解析水稻雄性生殖发育的整个调控网络。【方法】常规种植条件下,突变体ms7 (male sterile 7)与对照种植于浙江富阳和海南陵水,比较它们的育性及主要农艺性状差异,利用混池关联分析和图位克隆方法进行目标基因定位。【结果】整个生育期,突变体ms7生长速率与野生型一致,成熟期的株高、分蘖数、叶数、叶大小、穗长和每穗颖花数等性状与野生型相比也没有显著差异,但ms7结实率为0,表现为完全雄性不育,花药瘦小且颜色发白,半薄切片显示绒毡层降解推迟,花粉镜检呈染败。遗传分析表明花粉败育受单个隐性基因控制,定位于第7染色体上BSA11与YD7045之间1.17 Mb的范围内。【结论】本研究为水稻雄性不育基因ms7的克隆和功能研究打下了基础。     

The Difference in Sterility due to High Temperatures during the Flowering Period among Japonica-Rice Varieties

Summary

The objectives of this study are to compare the floret sterility induced by a high temperature given in the daytime during the flowering period among nine japonica rice varieties, and to clarify the mechanism of the high-temperature-induced sterility. Nine japonica rice varieties were subjected to 35.0, 37.5 or 40.0°C day- temperature conditions (1000-1600) for six consecutive days using sun-lit phytotrons, and the percentage of fertility, pollination and germinated pollen grains on the stigmas were examined. The temperature that caused 50% sterility varied with the variety, and a difference of approximately 3.0°C was observed between the most tolerant and susceptible varieties. Under the 37.5°C day-temperature condition, the percentage of florets with 10 or more germinated pollen grains was roughly coincident with the percent fertility, but under the 40° C day-temperature condition, it was higher than the percent fertility. Many of the florets with less than 10 germinated pollen grains had less than 20 total pollen grains on their stigmas under both temperature conditions. From these results, we concluded that sure pollination under high-temperature conditions is an important factor and that the high-temperature tolerance of the processes following pollen germination is also required for fertility under excessively high temperature conditions.     

Enclosed stigma contributes to higher spikelet fertility for rice(Oryza sativa L.) subjected to heat stress

With global warming, rice plants may be subjected to heat stress more regularly during the heatsensitive flowering stage, causing spikelet sterility and grain yield loss.Stigma exsertion is considered to increase pollen reception and promote female reproductive success.The aim of this study was to investigate the role of stigma exsertion on spikelet fertility at high temperatures.Five rice cultivars(Liangyoupeijiu, Shanyou 63, Huanghuazhan, Nagina 22, and IR64) with differing degrees of stigma exsertion were cultivated and exposed to high temperature at anthesis.Heat-tolerant cultivars did not always show a high percentage of spikelets with exserted stigmas, and vice versa.Irrespective of the presence of more pollen grains on exserted stigmas, spikelets with exserted stigmas did not show greater spikelet fertility than spikelets with fewer exserted stigmas or hidden stigmas under heat stress.GA3 application augmented the percentage of spikelets with exserted stigmas;however, it did not increase spikelet fertility under heat stress.Spikelet fertility of whole panicles was negatively correlated with the percentage of spikelets with exserted stigmas, but positively with that with hidden stigmas.Viability of the hidden stigmas was less reduced than that of exserted stigmas under heat stress, suggesting that hidden stigmas have an advantage in maintaining viability.Heat stress delayed anther dehiscence and reduced the viabilities of both exserted stigmas and pollens, thereby causing low spikelet fertility.Together, these results suggest that high spikelet fertility does not depend on stigma exsertion and that enclosed stigma generally contributes to higher spikelet fertility and heat tolerance under high-temperature conditions during flowering in rice.     

油菜异源花粉在柱头上萌发过程观察

以甘蓝型油菜为母本、白菜型油菜和芥菜型油菜为父本进行种间杂交．用扫描电镜和荧光显微镜对异源花粉在柱头上萌发及其花粉管进入柱头进行观察．结果表明，花粉管生长表现几种异常形态，即花粉管短，顶端球状膨大和棒状膨大、分杈，花粉管细长，均匀或不均匀扭曲．     

光敏核不育水稻育性转换条件及其生态适应性研究（英文）

 本文系统介绍了诱导光敏核不育水稻育性转换的光温作用模式的内容和意义,认为光敏核不育水稻育性转换的光周期诱导,只能在一定的温度范围内才存在。讨论了诱导光敏核不育水稻育性转换的光周期敏感期、光照长度、光照强度及曙暮光影响、光期暗期与光质的影响,分析了温度的作用、温度影响的敏感期与温度分解因子的影响;并对现有不育系的生态类型和适应性作了初步分析。     

Transgene Flow from Glufosinate-Resistant Rice to Improved and Weedy Rice in China

The development of transgenic rice with novel traits in China can increase rice productivity, but transgene flow to improved or weedy rice has become a major concern. We aimed to evaluate the potential maximum frequencies of transgene flow from glufosinate-resistant rice to improved rice cultivars and weedy rice. Treatments were arranged in randomized complete blocks with three replicates. Experiments were conducted between 2009 and 2010 at the Center for Environmental Safety Supervision and Inspection for Genetically Modified Plants, China National Rice Research Institute, Hangzhou, China. Glufosinate-resistant japonica rice 99-1 was the pollen donor. The pollen recipients were two inbred japonica rice （Chunjiang 016 and Xiushui 09）, two inbred indica rice （Zhongzu 14 and Zhongzao 22）, two indica hybrid rice （Zhongzheyou 1 and Guodao 1）, and one weedy indica rice （Taizhou weedy rice）. The offspring of recipients were planted in the field and sprayed with a commercial dose of glufosinate. Leaf tissues of survivors were analyzed by polymerase chain reaction to detect the presence of the transgene. The frequency of gene flow ranged from 0 to 0.488%. In 2009, the order of gene flow frequency was as follows： weedy rice 〉 Chunjiang 016 〉 Xiushui 09 and Zhongzu 14 〉 Guodao 1, Zhongzheyou 1 and Zhongzao 22. Gene flow frequencies were generally higher in 2009 than in 2010, but did not differ significantly among rice materials. Gene flow frequency was the highest in weedy rice followed by the inbred japonica rice. The risk of gene flow differed significantly between years and year-to-year variance could mask risk differences among pollen recipients. Gene flow was generally lesser in taller pollen recipients than in shorter ones, but plant height only accounted for about 30% of variation in gene flow. When flowering synchrony was maximized, as in this study, low frequencies of gene flow occurred from herbicide-resistant japonica rice to other cultivars and weedy rice. Averaged across years, the risk     

大气CO2浓度升高条件下移栽水稻与不同出苗时间稗草竞争的响应差异

通过田间试验,研究了FACE（开放式空气CO2浓度升高）条件下,移栽稻田水稻（Oryza sativa）与不同发生时期稗草（Echinochloa crusgalli）生长和竞争的响应差异。结果显示,FACE条件下水稻与稗草比例为1∶1时,水稻分蘖、生物量和产量均增加,而稗草则减少。水稻与移栽稻田本田萌生稗草的生物量比率增加181.7%、产量比率增加259.7%、茎蘖比率增加24.2%、有效分蘖比率增加186.3%,水稻竞争优势增加,而移栽稻田本田萌生稗草竞争能力下降。水稻与秧田萌生稗草的竞争关系变化趋势在营养生长期与上述情况相同,但进入生殖生长后水稻竞争优势不再显著,FACE条件下与秧田萌生稗草竞争的水稻最终生物量和产量分别增加22.7%和8.2%,但秧田萌生稗草的最终生物量和产量也增加了11.9%和9.5%。水稻与秧田萌生稗草的生物量比率、茎蘖比率和产量比率增加均不显著。 可见,田间大气CO2浓度升高条件下水稻与稗草的竞争关系发生的变化与稗草发生时期有重要关系。     

The Green Fence of Chinese Hibiscus (Hibiscus rosa-sinensis L.) Prevents Pollen Dispersal of Transgenic Rice (Oryza sativa)

Abstract

Transgene escape mediated by pollen dispersal is one of the preeminent concerns about genetically modified crops, including rice. In this study, the rice pollen donor of non-glutinous Oryza sativa ssp. japonica cv. ‘Tainung 67’ [TNG 67] had a greater potential of pollen flow, which was shown by the greater quantity, germination rate, and viability of pollen, than the other rice pollen donor of transgenic AAN. The pollen-mediated gene flow was detected by the frequency of outcrossed seeds in a “checker-board pattern” and alternating row arrangement of rice pollen donor (TNG 67 or AAN) and pollen recipient (glutinous rice ‘TNG 73’) in the fields. We conducted field experiments to assess rice pollen dispersal with or without a “green” fence of Chinese hibiscus, Hibiscus rosa-sinensis L., of approximately 2 m in height and 0.6 m in thickness. Without a green fence, the outcrossing rate of TNG 73 seeds decreased with increasing distance from the pollen donor, from 1.68% at 1 m to 0.01% at 35 m, with no outcrossing beyond 40 m. The outcrossing rate varied with the direction of pollen donor, from 0.05% in the northeastern direction to 0.78% in the southern direction, which was caused by prevailing wind direction. With the green fence, no seeds of TNG 73 were outcrossed. Since a buffer zone of at least 40 m is needed to prevent outcrossing of rice by pollen dispersal in an open field, planting of Chinese hibiscus around the rice field as a green fence would be an effective measure for preventing transgene escape mediated by pollen flow.