Controlling the release of pollen from genetically modified maize and increasing its grain yield by growing mixtures of male-sterile and male-fertile plants

There is public concern about the consequences of pollen dispersal from genetically modified (GM) crops. There lease of viable pollen from GM maize can be controlled by growing mixtures of cytoplasmic male-sterile plants and male-fertile non-transformed pollinator plants. Our experiments indicate that such associations can bring about grain yields as high or even higher than those produced by pure male-fertile maize crops, especially when the male-sterile component is pollinated non-isogenically. The grain yield benefits from cytoplasmic malesterility and xenia as well as the fact that seed of male-sterile varieties can be produced cheaply and reliably in large quantities would facilitate the implementation of the proposed system in agricultural practice. This revised version was published online in July 2006 with corrections to the Cover Date.     

转AtHDG11基因玉米的获得及抗旱性鉴定

培育抗除草剂和抗旱的转基因玉米种质,通过超声波辅助农杆菌介导花粉非组培法将拟南芥抗旱基因At HDG11导入玉米自交系昌7-2中,获得转基因植株。通过逐代除草剂筛选、分子检测对获得的转基因植株进行检测,获得5株转基因株系。在此基础上,以非转基因自交系昌7-2为对照,对5个转基因株系进行抗旱性鉴测;对苗期和十叶期玉米植株进行干旱胁迫处理,测定转基因植株生理活性物质和光合参数。结果表明,在干旱胁迫条件下,转基因玉米叶片的Pro含量高于非转基因玉米,MDA含量低于非转基因玉米;不论在正常生长条件下还是干旱条件下,转基因株系的光合速率和水分利用率都明显高于对照植株;而转基因株系的蒸腾速率和气孔导度则明显低于对照植株。综合室内与田间的抗旱检测结果表明,超声波辅助农杆菌介导花粉的非组培转基因方法在玉米上是可行的。将拟南芥HDG11基因导入玉米,同样可以有效地提高玉米的抗旱性,为玉米的抗旱育种提供新的种质资源和新的转基因方法。     

Effect of pollinator on haploid production in durum wheat crossed with maize and pearl millet

The aim of this work was to analyse the influence of the male parent on the production of embryos and haploid plants in durum wheat crossed with maize and pearl millet, to find a proper trait to identify the most efficient pollinators and to evaluate the mixtures of pollen. Two genotypes of durum wheat, low and high responding, were crossed with eight pollen samples: (i) three maize hybrids, (ii) three pearl millet inbred lines, (iii) a mixture of maize pollen and (iv) another mixture of pearl millet pollen. No significant differences on embryos and haploid plant production were observed among the four samples of maize pollen, but there were clear genotypic differences for the production of haploids between genotypes of pearl millet. The best pearl millet genotype produced significantly more haploid plants than the other two and the mixture of pollen. There was no correlation between the production of embryos and haploid plants. Therefore, the production of haploid plants must be the criterion to identify superior pollinators. In addition, a mixture of pollen is inappropriate except when using genotypes previously identified as good pollinators.     

水分胁迫对玉米花粉活力和花受精能力的影响

抽穗期的玉米植株经不同强度水分胁迫处理后,花粉和花丝中ＳＯＤ活性下降,膜脂过氧化作用增强,对生物膜系统产生直接伤害,电解质外渗率增加,电镜下观察到线粒体和细胞质膜破坏,胞间连丝膨大变形,甚至断裂,粗糙内质网卷曲成空心筒状,甚至部分解体,导致代谢紊乱,物质运输受阻,影响花粉和花丝发育,降低雌穗花丝受精能力,减少穗粒数,从而造成减产。耐旱性不同的玉米,其花粉和花丝活力及花丝受精能力受水分胁迫的影响不同     

Pollen-mediated gene flow in an autogamous crop: Foxtail millet (Setavia italica)

Gene How is a key concern associated with the eonlamination of seed multiplication iields and the use of transgenic crops. The release of herbicide-resistant germplasnis and the use of male-sterile varieties make foxtail millet (Sviaria ilalicii) an appropriate material to inves-tigate this concern. Pollen dispersal from pollen donor sources and gene tlow in fertile and male-slerile varieties of fortail millet were investigated in experiments in China and France. Although foxtail millel is mainly autogamous (less than 2% oiitcrossing), pollen from a 5m diameter plot could fertilize a tertile variety up to 24 ni away, and a male-sterile variety up to 40m. Negative exponential and WeibuU fnnctions were used to tit the relationships between percentage and number i.A iubrids per cm of ear with distance from the pollen source. Wind velocity and direction resulted in variations of gene How intensity, but did nol change liie slope ofthe regression equations. Pollen competition for available target ovules was observed up to 2 m away from a 1 m diameter pollen source. The percentage of pollen grains that fertilized ovules outside the pollen donor source was 1.4% of the total pollen grain produetion. For every iflO pollen grains released under lieid conditions 1.5 seeds were produced. The probability funetion of pollen dispersal for one donor plant was calculated. It allowed predictions of gene How accord-ing to lield shapes, estimations of isolation distances to preserve /ariety purity against pollen contamination, and the fornuiiation of efficient male/female ratios Ibr hybrid seed production.     

High crop barrier reduces gene flow from transgenic to conventional maize in large fields

To test the effectiveness of high crop barriers in reducing gene flow, we conducted experiments in two large fields (around 750 m × 750 m) using yellow transgenic maize and white conventional maize in the growth season of 2013, with Sorghum as a high crop barrier in a shape of 5 m zone. Sorghum barrier affected pollen load at different directions, decreasing pollen number in further locations. It also decreased the cross-pollination incidence between transgenic and non-transgenic maize, with a average rate of 9.35% in the open site and 1.04% in the Sorghum site. But Sorghum barrier had little effect on the maximum distance of pollen flow and cross-pollination, which depends on wind direction and speed. The maximum distance of gene flow from transgenic to conventional maize was 300 m in the open site and 350 m in the Sorghum site. High crop barrier could be proposed as an effective method to reduce the frequency of gene flow from transgenic to conventional crops and to regulate their coexistence.     

Wheat pollen dispersal under semiarid field conditions: potential outcrossing with Triticum aestivum and Triticum turgidum

Isolation distance is the main barrier to crop-to-crop gene-flow. A 3-year study assessed the maximum potential outcrossing under field conditions between two wheat cultivars (Triticum aestivum L.) and between wheat and durum wheat (Triticum turgidum L. var. durum). Outcrossing was measured by seed set on emasculated recipient plants placed at four sides with different distances from a 3 m × 3 m T. aestivum (cultivar Chinese Spring) pollen source. Frequencies of seed set at 0 m distance were 45% (37–56%) for T. aestivum cultivars and 18% (5–30%) with T. turgidum. These values agree with hybridization in non-limiting pollen conditions measured by manual crosses in greenhouse. The number of pollen grains and the outcrossing frequencies decreased at increasing distances influenced by the prevailing wind direction. Under semiarid conditions of this assay, viable pollen was found 14 m from the pollen source, with a maximum distance of 8 m at which cross-pollination decreases below 1%. Ambient conditions affect pollen viability, hybridization and pollen dispersal. Data presented in this paper emphasize the major role played by environmental conditions in outcrossing. Data obtained in one area may therefore not coincide with the prevailing situation in different locations and climates.     

Ectopic expression of a maize pollen specific gene, zm401, results in aberrant anther development in tobacco

Our previous study shows that the maize zm401 cDNA is specifically expressed in maize pollens. This evidence suggests that zm401 likely functions in pollen growth and/or development. To confirm its possible involvement in pollen development, the full length cDNA of zm40l was ectopically expressed in tobacco plants under the control of a pollen specific promoter ZM13 (from maize). RT-PCR amplification demonstrated that the ZM13-driven zm401 gene was spatially expressed in tobacco pollens. It was found that all transgenic tobacco plants expressing zm401 showed various levels of sterility, ranging from abortive flower development to male sterility. Further analyses on anther development of transgenic plants indicated multiple abnormalities in the late stages of anther development. These abnormalities include lagged degradation of the tapetum and connective tissue, failed deposition of fibrous bands in endothecium cells, and aborted pollen grain development. These results strongly suggest that zm401 plays an essential role in anther development. However the exact functions of zm401 is still unclear, and further analysis of zm401 is required to determine the exact mechanism involved in anther development.     

Assessing the risks of wind pollination from fields of genetically modified Brassica napus ssp. oleifera

Summary Intensive research over the past 10 years has produced many genetically-mofified lines of oilseed rape with market potential. Assessment of these lines in statutory trials prior to their release as cultivars is necessary, owing to concern over the likelihood of transgene escape from such crops. Here, we examine the movement of airborne pollen grains from oilseed rape fields and assess their capacity for long-range geneflow.Pollen dispersal from isolated rape fields was monitored over two seasons and related to the distribution of fields and feral (domesticated plants growing outside cultivations) populations of the crop in Tayside and North East Fife regions of Scotland. Airborne pollen density declined with distance and at 360 m was 10% of that at the field margin. Pollen counts of 0–22 pollen grains m³ were observed 1.5 km from source fields and apparently were sufficient in number to allow seed set on emasculated bait plants. Oilseed rape pollen has greater capacity for long-range dispersal than had been suggested by small-scale field trials. Mean separation of oilseed rape fields in the survey area was 410 m and the mean distance from feral populations to commercial fields was 700 m. Sixty percent of feral populations with more than 10 plants occurred downwind and within 2 km of an oilseed rape field. Provided that the flowering biology of genetically-modified oilseed rape does not differ from the conventional crop, these data suggest that transgene movement to non genetically-modified fields or feral populations is likely following commercial release.     

The role of pollen and stigma in determining the level of partial self-compatibility in brussels sprouts

Summary Brussels sprouts inbred plants, homozygous for the same recessive S-allele, were intercrossed in all combinations and the number of pollen tubes penetrating the stigmas of such crosses measured. The results of five such experiments showed that pollen as well as stigma is significant in determining the number of pollen tubes counted. It was also found that the capacity of a plant's stigmas to permit pollen tube penetration was independent of the capacity of its pollen to penetrate other plants' stigmas. Variation in pollen tube production between pollens of the plants tested was often greater than that between their stigmas.     

温度对白魔芋花粉功能及自交结实的影响

研究温度对白魔芋散粉、花粉萌发、活力保持及自交结实的影响,以离体萌发法测定花粉萌发率,以培养箱法测定恒温自交结实率,以野外套袋法测定自然自交结实率。结果显示:最佳散粉温度22~26℃,18℃低温下可正常散粉,而34~38℃高温下80%以上花株花粉囊无法正常开裂;高浓度钙可加快花粉管的生长;花粉在18、22、26、30、34、38℃下的萌发率分别为8.78%、33.17%、54.90%、38.83%、29.10%、6.98%;花粉在38、34、30、26、22、18℃贮藏下分别在4、4、12、12、25、55 h内丧失活性。恒温培养中仅26℃下有4.35%的自交存活率,野外套袋中日平均温度为23.7℃时有2.94%的自交存活率。白魔芋的散粉培养及离体萌发最佳温度为26℃,温度过高或过低均影响授粉结实率。及时采集花粉、控制温度、调节湿度、适当灌溉均能提高授粉结实率。     

热胁迫下对玉米自交系育性的鉴定

在山西运城市7-8月份气温偏高的特殊气候环境下,对120份玉米自交系材料进行了雄花育性鉴定。结果表明,高温条件下,材料间的雄花育性存在着明显的差异;不育性表现方式有:无花粉粒、花药吐不出、花药不散粉。模拟玉米螟为害雄花节打孔试验表明,即使玉米植株能得到充足的水分供应,雄花内部的水分胁迫是影响花粉正常散出的主要因素之一。     

Dicamba and growth condition effects on doubled haploid production in durum wheat crossed with maize

Doubled haploid plants are useful in genetic studies and plant breeding, but a consistent and satisfactory frequency of production has been difficult to achieve in durum wheat. Triticum turgidum L., using the maize pollen method. The objective of this study was to develop an objective method of producing doubled haploids in durum wheat. Plant growing and handling conditions, aspects of hormone treatments, wheat genotype and pollen source were considered. The number of caryopses, embryos, haploids, doubled plants and doubled plants that set seed were measured. Although growth conditions, pollen source, method of handling plants and wheat genotype are important considerations, the type of hormone was found to be most significant in the production of doubled haploid plants. When 50mg/l dicamba was substituted for 100 mg/l 2,4‐D the number of doubled haploids per spike increased from 0.2 for the best 2,4‐D treatment to 1.3 for the dicamba treatment. This increased frequency was largely attributed to an increase in the number of caryopses generated for each spike emasculated and from an increased frequency of germination of embryos to haploid plantlets. The best production of caryopses was 0.41 caryopses per florest with 2,4‐D. The best production of haploids per 100 florets was 12 with dicamba and 1.65 with 2,4‐D. The frequency of one doubled haploid per emasculated spike through the use of dicamba is a practical level for generating populations for genetic studies.     

New maize × Tripsacum hybrids for maize improvement

Summary Approximately 27 000 maize x tripsacum pollinations were made and 19 hybrids were identified. Fourteen of these hybrids were of the classical type, which possesses the full gametic complement of chromosomes from each parent. These hybrids are more like tripsacum than maize. They are perennial, tiller profusely, and begin to flower in approximately two years from germination. So far, none of these hybrids has produced pollen and all backcrosses have been unsuccessful.Five of the 19 hybrids were not of the classical type, and possessed a variable number of chromosomes in cells of the same root tip. Most cells examined contained 20 maize and no tripsacum chromosomes. Other cells contained 20 maize chromosomes and up to eight of tripsacum. However, in at least three hybrids some cells contained a total of 20 chromosomes, of which up to four were tripsacum substitution chromosomes. These five non-classical hybrids are more maize-like, and are annual. Most have flowered within three months of germination. Although they have produced no viable pollen, four plants have set seed after backcrossing to maize.The production of F₁ hybrids is discussed with view to their subsequent assessment and utilization for maize improvement.     

Effects of Nitrogen Application Time on Ear Components of Maize

Pot and field experiments were conducted with two varieties of maize to study the effects of nitrogen applied in different doses. The total amount of N applied was same in all variants of one experiment. The development of maize plants varied due to variety and N-doses. In pot experiment no N application before sowing or in the begin of the ear differentiation reduced total number of spikelets, spikelet rows/ear, length of ear, kernel yield as well as plant height. In field experiments an amount of 30 kg N/ha in the begin of ear formation was sufficient for maximum increase in almost all yield characteristics. Higher doses than that were not effective to promote these parameters further.     

基于RNA-seq技术的玉米SNP和InDel标记分析

高温已对玉米(Zea mays L.)生产造成了巨大危害。为了给耐高温胁迫玉米品种选育提供可用的分子遗传标记,本研究以耐高温和高温敏感的两个玉米品种为实验材料,通过花粉总RNA提取、mRNA纯化、反转录、转录组文库构建及测序等工作,对获得的转录组数据进行SNP和InDel分子标记分析。结果显示:高质量碱基数据量平均为6.69 Gb,占原始数据比例平均达94.71%;在SNP分子标记中,Homo位点平均为19450个,Hete位点平均为23598个,转换类型共52337个,颠换类型共27868个;在InDel分子标记中,Homo位点平均为1023个,Hete位点平均为1736个;SNP/InDel位点在外显子区分布最多,达到总量的58.02%,其次是基因间隔区、3'非翻译区和5'非翻译区,其余区域位点占总量比例均低于5%;SNP/InDel造成同义单核苷酸突变比例最高,平均值为60.69%,其次为非同义单核苷酸突变,平均值为36.72%,非移码替换突变为平均值1.64%,其余3种均不到1.00%;在SNP和InDel标记位点基因功能注释中,NR数据库注释最多,超过8000个,其次为KEGG和eggNOG;注释到GO和Swissprot数据库的相对较少。本研究结果为进一步标记辅助耐高温玉米品种选育提供了分子基础,也为其它植物的类似研究提供了参考。     

孕穗期高温逆境对玉米杂交种的影响分析

为了筛选和分析耐高温逆境能力强的夏玉米品种,利用智能温室在玉米孕穗期模拟高温气象,研究分析孕穗期高温逆境对10个夏玉米杂交种的植株性状、穗部性状、花期、可育率和理论产量等指标的影响。结果表明,高温逆境使孕穗期玉米的株高和穗位高变高,抽雄期、吐丝期和散粉期滞后,抽雄至散粉间隔期缩短,雌雄间隔期延长,穗长、穗粗、穗行数、行粒数、穗粒数、穗粒重、可育率和理论产量下降。通过对不同玉米品种在孕穗期高温逆境下的性状表现及其变化的差异分析和聚类分析,试验品种的耐高温热害能力由强到弱表现为：漯玉336>漯玉16>阳光98>郑单958>豫丰3358>先玉335>阳光99>漯青19>漯玉172>漯玉171。     

Natural outcrossing in common buckwheat

Common buckwheat is an obligate cross pollinating crop because of its sporophytic self-incompatibility system. Therefore a study to assess the extent of natural outcrossing was undertaken for two years at Morden and Portage la Prairie, Manitoba using the semi-dwarf character, which is due to a homozygous genotype with regard to a single recessive gene, as a marker. The semi-dwarf genotype was grown in 100 m rows running in four directions at 90 ° from a central 36 m² plot of a normal, tall variety. At maturity, seed samples were taken from the semi-dwarf population at designated intervals. At least 200 F₁ seeds from each sample were grown in the greenhouse to the 3-4 leaf stage and the proportion of tall plants was used to determine the percentage of outcrossing. Approximately 50% outcrossing occurred where the semi-dwarf plants were immediately next to the normal plants and then the proportion decreased with increasing distance. Although outcrossing occurred throughout the distance tested, there were no significant differences from 12 m onward from the pollen source. Lack of directional influence in outcrossing and a sharp reduction in the proportion of tall plants within a 3 m distance from the pollen source suggested that wind was not a major factor in dispersing the buckwheat pollen over long distances. Although a substantial cross pollination was limited to a few metres from the pollen source, nearly 1% outcrossing occurred even at 100 m. Since the distance in this experiment was limited to 100 m from the pollen source, the present results do not allow us to recommend the minimum isolation distance required to meet certain standards for buckwheat seed production. This revised version was published online in July 2006 with corrections to the Cover Date.     

玉米雄性不育突变体mi-ms-3的遗传分析及分子鉴定

Maize is one of the best crops in the utilization of heterosis. Male sterile lines are important germplasms for the hybrids production. A male sterile mutant named mi-ms-3 was obtained by screening in a mutator insertion library. The number of male anthers in tassel decreased and not exserted. There were few anthers with only two pollen sacs in the mutant tassels, and some of the anthers were degenerated to membranous and formed filaments at their ends. Although pollens in the anthers could be stained by I₂-KI, pollen shedding was abnormal and the number of pollen grains decreased. The number of silks in the ear of the mutant increased, and there was a sterile grain on both sides of the maturated kernel. Fertility of F₁ plants, which were obtained by hybridization between mi-ms-3 and maize inbred Mo17, was normal. Genetic analysis of F₂ population showed that the mutant phenotype was controlled by a recessive gene. The candidate gene was preliminarily mapped on the long arm of chromosome 3 by BSA and it was located between a SSR marker and an Indel marker with a distance of 1.5 cM. There are 21 candidate genes in this region. It was finally found that the insertion mutation of Mu transposon occurred at 30 bp upstream of the coding region of zm00001d042618 (zmm16) by transponson tagging and sequencing analysis. The results showed that mi-ms-3 was a new allele of sts1, which caused by a single base mutation in the coding region. RT-PCR analysis indicated that the expression of zmm16 in the mutant was decreased. The identification of the new allelic mutant of sts1 in this study would provide new materials for the study of flower development and hybrid seed production.     

Characteristics of Pollen from Transgenic Lines of Apple Carrying the Exogenous CpTI Gene

It is fundamental for gene transformation and ecosystem hazard evaluation to study the pollen characteristics of transgenic plants. In this research,the characteristics of pollen from 7-or 8-year-old transgenic apple plants carrying an exogenous Cp TI gene were analyzed. The results showed that there was no significant difference in terms of size,morphology,or exine ornamentation between the pollen of the transgenic plants and the non-transgenic control. However,the transgenic plants had more abnormal pollen grains. Of the 13 transgenic lines tested,12 had a significantly lower amount of pollen and six exhibited a significantly lower germination rate when cultured in vitro. The pollen viability of three transgenic lines was determined,with two showing significantly lower viability than the control. The transgenic Gala apple pollen grains germinated normally via controlled pollination on Fuji apple stigmas. However,the pollen tubes extended relatively slowly during the middle and late development stages,and another 8 h were needed to reach the ovules compared with the control. The gibberellic acid concentration in transgenic Gala apple flowers was lower than in the non-transgenic control during all development stages tested. The abscisic acid concentration in the transgenic flowers was lower during the pink stage,and higher during the ball and fully open stages. Microscopic observation of the anther structure showed no difference. The tapetum of the pollen sac wall in transgenic plants decomposed late and affected pollen grain development,which could be one of the reasons for the lower number of pollen grains and poor viability in the transgenic plants.