Comparison between Two Methods for Detecting Gene Flow and Analysis on the Influencing Factors

[Objective] The aim was to test the consistency between two commonly used methods PCR detection and protein detection for detecting gene flow,and investigate the factors influencing the consistency.[Method] 1 769 samples of three varieties under three treatments（wind,bee and control）were detected with both methods.[Result] There was phenomenon in field that Bt gene was transferred into F1 generations but couldn＇t express,that meant,the result of PCR detection was not consistent with that of protein detection.By comparing environmental factors,it was proved that wind and bee treatments didn＇t significantly affect the expression of gene flow.However,the Bt gene expression rate in bee treatment was higher than that in wind treatment.[Conclusion] The paper will provide reference for accurate detection of gene flow.

Gene flow from transgenic roundup-ready soybean to wild soybean

A study was conducted in the field of the Institute of Vegetable Crops, Jiangsu province from July 2000 to August 2003. The transgenic roundup-ready soybean was sown in the middle of the field in a circular manner for 5 circles, with the distance of 3 m, from one circle to another. Then the wild soybean was planted in plots as the rays of the circles in 8 directions (N, E, W, S, NE, NW, SE and SW), spaced every 5 m until 50 m. Each plot comprised 25 plants. In the second year, the wild soybean seeds from the first year were planted in the field together with the original wild soybean as check. Before flowering time, high concentrations of roundups (about 4-5 times of the normal dose) were sprayed on the plants and the surviving plants were identified. The leaves were taken to the lab for DNA extraction to determine the unique DNA for roundup-ready soybean (CTAB method). About 2% of the plants survived, but some leaves were yellow. One plant of wild soybean was found to have the roundup-ready gene from the original roundup-ready soybean. The other surviving wild soybeans should also had some fragments of the roundup tolerance gene. However, the DNA bands were not very clear in the PCR map.

High gene flows promote close genetic relationship among finewool sheep populations(Ovis aries) in China

The aim of our present study was to construct genetic structure and relationships among Chinese fine-wool sheep breeds. 46 individuals from 25 breeds or strains were genotyped based on the Illumina Ovine 50K SNP array. Meanwhile, genetic variations among 482 individuals from 9 populations were genotyped with 10 microsatellites. In this study, we found high genetic polymorphisms for the microsatellites, while 7 loci in the Chinese superfine Merino strain （Xinjiang types） （CMS） and 5 loci in Gansu alpine superfine-wool sheep strain （GSS） groups were found deviated from Hardy-Weinberg equilibrium （HWE）. Genetic drift FsT=0.019 （P〈0.001） and high gene flows were detected in all the 7 fine-wool sheep populations. Phylogenetic analysis showed fine-wool sheep populations were clustered in a group independent from the Chinese indigenous breeds such that the 7 fine-wool sheep clustered distinct from Liangshan semifine-wool sheep （LS） and Hu sheep （HY） reflected by different population differentiation analyses. Overall, our findings suggested that all fine-wool sheep populations have close genetic relationship, which is consistent with their breeding progress. These populations, therefore, can be regarded as open-breeding populations with high levels of gene flows. Furthermore, the two superfine-wool strains, viz., CMS and GSS, might be formed by strong artificial selection and with frequent introduction of Australian Merino. Our results can assist in breeding of superfine-wool sheep and provide guidance for the cultivation of new fine-wool sheep breeds with different breeding objectives.

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 of gene flow to weedy rice was higher than that of improved rice and hybrids. Greater resources must be dedicated to the management of remnant weedy rice in fields planted with herbicide-resistant rice, and to prevent the evolution of resistant weedy rice populations.

Genome-wide analysis reveals signatures of complex introgressive gene flow in macaques(genus Macaca)

The genus Macaca serves as an ideal research model for speciation and introgressive gene flow due to its short period of diversification(about five million years ago)and rapid radiation of constituent species.To understand evolutionary gene flow in macaques,we sequenced four whole genomes(two M.arctoides and two M.thibetana)and combined them with publicly available macaque genome data for genome-wide analyses.We analyzed 14 individuals from nine Macaca species covering all Asian macaque species groups and detected extensive gene flow signals,with the strongest signals between the fascicularis and silenus species groups.Notably,we detected bidirectional gene flow between M.fascicularis and M.nemestrina.The estimated proportion of the genome inherited via gene flow between the two species was 6.19%.However,the introgression signals found among studied island species,such as Sulawesi macaques and M.fuscata,and other species were largely attributed to the genomic similarity of closely related species or ancestral introgression.Furthermore,gene flow signals varied in individuals of the same species(M.arctoides,M.fascicularis,M.mulatta,M.nemestrina and M.thibetana),suggesting very recent gene flow after the populations split.Pairwise sequentially Markovian coalescence(PSMC)analysis showed all macaques experienced a bottleneck five million years ago,after which different species exhibited different fluctuations in demographic history trajectories,implying they have experienced complicated environmental variation and climate change.These results should help improve our understanding of the complicated evolutionary history of macaques,particularly introgressive gene flow.

Multilocus phylogeography and ecological niche modeling suggest speciation with gene flow between the two Bamboo Partridges

Background:Understanding how species diversify is a long-standing question in biology.The allopatric speciation model is a classic hypothesis to explain the speciation process.This model supposes that there is no gene flow during the divergence process of geographically isolated populations.On the contrary,the speciation with gene flow model supposes that gene flow does occur during the speciation process.Whether allopatric species have gene flow during the speciation process is still an open question.Methods:We used the genetic information from 31 loci of 24 Chinese Bamboo Partridges(Bambusicola thoracicus)and 23 Taiwan Bamboo Partridges(B.sonorivox)to infer the gene flow model of the two species,using the approxi-mate Bayesian computation(ABC)model.The ecological niche model was used to infer the paleo-distribution during the glacial period.We also tested whether the two species had a conserved ecological niche by means of a back-ground similarity test.Results:The genetic data suggested that the post-divergence gene flow between the two species was terminated before the mid-Pleistocene.Furthermore,our ecological niche modeling suggested that their ecological niches were highly conserved,and that they shared an overlapping potential distribution range in the last glacial maximum.Conclusions:The allopatric speciation model cannot explain the speciation process of the two Bamboo Partridges.The results of this study supported a scenario in which speciation with gene flow occurring between the allopatric species and have contributed to our understanding of the speciation process.

Assessment of Gene Flow Through Detection of SexualCompatibility Between Transgenic Rice with barand Echinochloa crusgalli var.mitis

The possibility of gene flow between two varieties of transgenic rice with bar gene (Y0003 and 99-t) (male) and barnyard grass(Echinochloa crusgalli var. mitis ) (female) was studied by means of reproductive biology. The germination and growth of rice pollen grains on barnyard grass stigmas at 30 min, and 1-4 h after crossing by hand were observed with an optical microscope. The results were compared with the germination and growth of barnyard grass pollen grains at the corresponding time after self-pollination. The results showed that germination and growth of the pollen grains of the two varieties were similar on barnyard grass stigmas, but differed significantly from self-pollination of barnyard grass. Pollen grains germinated and pollen tubes penetrated stigmas normally, and the number of pollen grains being condensing or releasing their inclusions or having released them increased with the time after self-pollination. Pollen grains of transgenic rice on the stigmas of barnyard grass couldn't germinate or grow normally after crossing, neither could they penetrate the stigmas of barnyard grass. Therefore, it could be concluded that the sexual incompatibility between transgenic rice with bar gene and barnyard grass is due to the rice pollen being unable to penetrate the stigma of barnyard grass. Further proof of incompatibility lies in the fact that the emasculated barnyard grass pollinated with the rice pollen grains could not seed.

Evolutionary history of a desert perennial Arnebia szechenyi(Boraginaceae):Intraspecific divergence,regional expansion and asymmetric gene flow

The complex interactions of historical,geological and climatic events on plant evolution have been an important research focus for many years.However,the role of desert formation and expansion in shaping the genetic structures and demographic histories of plants occurring in arid areas has not been well explored.In the present study,we investigated the phylogeography of Arnebia szechenyi,a desert herb showing a near-circular distribution surrounding the Tengger Desert in Northwest China.We measured genetic diversity of populations using three maternally inherited chloroplast DNA(cpDNA)fragments and seven bi-paternally inherited nuclear DNA(nDNA)loci that were sequenced from individuals collected from 16 natural populations across its range and modelled current and historical potential habitats of the species.Our data indicated a considerably high level of genetic variation within A.szechenyi and noteworthy asymmetry in historical migration from the east to the west.Moreover,two nuclear genetic groups of populations were revealed,corresponding to the two geographic regions separated by the Tengger Desert.However,analysis of cpDNA data did not show significant geographic structure.The most plausible explanation for the discrepancy between our findings based on cpDNA and nDNA data is that A.szechenyi populations experienced long periods of geographic isolation followed by range expansion,which would have promoted generalized recombination of the nuclear genome.Our findings further highlight the important role that the Tengger Desert,together with the Helan Mountains,has played in the evolution of desert plants and the preservation of biodiversity in arid Northwest China.

Gene flow的溯源及其在生物安全和进化研究中的意义

尽管gene flow源于群体遗传学和进化生物学,但已成为环境生物安全文献中常见的科学术语。花粉介导的gene flow在自然界中广泛存在并对物种和群体的进化有着特殊重要的意义。随着转基因生物技术的快速发展和转基因作物在全球范围内的广泛种植,转基因随gene flow发生逃逸及其可能带来的潜在生态进化影响已经成为环境生物安全评价和研究的重要内容,备受全球广泛关注,gene flow这个术语在我国也被频繁引用。但是,gene flow的中文术语在我国各种文献资料中存在着十多种翻译版本,这些不同的翻译版本形式不同且内容略有差异,容易给环境生物安全问题的理解和研究造成不必要的混乱。本文对gene flow的概念及其内涵进行了回顾,并对不同形式的gene flow术语在国内外相关研究领域中使用的历史溯源进行了阐述。笔者建议使用"基因流"作为gene flow在中文应用中的统一术语,这也最接近群体遗传学和进化生物学等相关著作中gene flow的原意。基于此,对基因流在转基因逃逸及其相关的环境生物安全评价以及群体遗传学和进化生物学研究方面的理论和应用意义进行了讨论。

Parapatric speciation with recurrent gene flow of two sexual dichromatic pheasants

Understanding speciation has long been a fundamental goal of evolutionary biology.It is widely accepted that speciation requires an interruption of gene flow to generate strong reproductive isolation between species.The mechanism of how speciation in sexually dichromatic species operates in the face of gene flow remains an open question.Two species in the genus Chrysolophus,the Golden Pheasant(C.pictus)and Lady Amherst’s Pheasant(C.amherstiae),both of which exhibit significant plumage dichromatism,are currently parapatric in southwestern China with several hybrid recordings in field.In this study,we estimated the pattern of gene flow during the speciation of the two pheasants using the Approximate Bayesian Computation(ABC)method based on data from multiple genes.Using a newly assembled de novo genome of Lady Amherst’s Pheasant and resequencing of widely distributed individuals,we reconstructed the demographic history of the two pheasants by the PSMC(pairwise sequentially Markovian coalescent)method.The results provide clear evidence that the gene flow between the two pheasants was consistent with the predictions of the isolation with migration model during divergence,indicating that there was long-term gene flow after the initial divergence(ca.2.2 million years ago).The data further support the occurrence of secondary contact between the parapatric populations since around 30 kya with recurrent gene flow to the present,a pattern that may have been induced by the population expansion of the Golden Pheasant in the late Pleistocene.The results of the study support the scenario of speciation between the Golden Pheasant and Lady Amherst’s Pheasant with cycles of mixing-isolation-mixing,possibly due to the dynamics of geographical context in the late Pleistocene.The two species provide a good research system as an evolutionary model for testing reinforcement selection in speciation.

Presence of G84E Allelic Heterogeneity of the European Prostate Cancer SNP Mutation of HOX13B-G84E Associated with the European R-Haplogroup of Y-Chromosome and Absence of Gene Flow into Moroccans Patients

SNP mutations in the HOXB13 gene associated with prostate cancer were determined in Moroccans prostate cancer patients (PCa). All PCa SNP mutations were new and belong to the SNP point-mutations located on the stop codon of HOXB13 exon 1 and 2 located in chromosome 17. The five mutations and their frequencies were as follows: rs1197613952 (12%), rs1597934612 (4%), rs1597933874 (4%), rs1597933837 (4%) and rs867793282 (4%). The European HOXB13-G84E (rs138213197) PCa mutation was not detected among Moroccan patients. The Y-chromosome genealogical haplotypes of the Western European (R1b1b2-M2G9) and the Eastern European (R191a-M-17) were not observed in Moroccans PCa patients. The patients have their own haplotypes E1b1 and J with a frequency of 55 and 35%, respectively. The results of the SNP mutations in the HOXB13, the absence of the HOXB13-G84E of the European in the Moroccans PCa patients, the absence of the European-lineage haplogroups (R1a1a-M17 and R1b1b2-M269) and the presence of E1b1b and J in Moroccans PCa patients would clearly indicate the absence of gene flow from European to Moroccans gene pool.

Theoretical Basis of Gene Splitting Technique and Its Application in the Control of Transgene Flow

Transgenic safety issues cause more and more controversies with the planting area of transgenie crops increased year by year. Gene flow from transgenie crops to wild relatives through pollen dispersal is one of the focus problems. Gene splitting technique provides a new strategy for the control of transgene flow by bio-logical containment. The construction of gene splitting technique is based on protein trans-splicing mediated by intein. Currently, it has been proved in Arabidopsis, tabaoco, wheat, etc. that active and functional proteins can be reassembled by intein mediated protein trans-splicing after gene splitting, which provides theoretical basis and experimental supporting for the limit of transgene flow by gene splitting technique. The theoretical basis of gene splitting technique and research progresses of its application on the control of transgene flow were reviewed in this paper.

Simulating the Gene Flow of Genetically Modified Maize in Taiwan

A field experiment was conducted in Taiwan to measure the cross-pollination (CP) rate of maize pollen recipients from pollen sources using phenotypic marker and to determine the isolation dis- tance between the 2 maize varieties. A waxy variety (Black Pearl) with purple kernels simulated the genetically modified (GM) pollen donor, and another waxy variety (White Pearl) with white kernels simulated the non-GM recipient. For the first crop, the total area was approximately1.5 hawith a pollen source and recipient acreage ratio of approximately 1:32. For the second crop, the total area was approximately1.83 hawith a ratio of approximately 1:17.3. The source fields were surrounded by the recipient fields for 2 crop seasons. The results showed that the rate of CP was <0.05% beyond15 mupwind and84.8 mdownwind in all crop seasons. The CP rate was below 5% at a distance of10min the downwind direction. A sample with 0.24% CP was recorded at107.3 mdownwind;however, the CP rate was 0% at68 mupwind. Three empirical models were used, that is, exponential, log/log and log/log, and a simplified Gaussian Plume model, to examine the relationship between the CP rates and the source-field distances. All of the models were appropriate for predicting CP rates, and the Gaussian Plume model performed better compared to the empirical models. The results show that it is possible to control CP from foreign pollen by using an appropriate isolation distance.

Gene deletor： a new tool to address gene flow and food safety concerns over transgenic crop plants

Environmental and food safety concerns over transgenic plants have hampered commercial applications of transgenic plant technology worldwide. A recently developed transgene deletion technology, named gene deletor technology, may be used to eliminate all transgenes from pollen, seeds, fruits or other organs when functions of transgenes are no longer needed or their presence may cause concerns. In this review, I will briefly describe the principle of the gene deletor technology with major supporting experimental data. I will also explain main characteristics and requirements of the gene deletor technology. Finally, I will discuss the gene deletor technology in the context of how it may be used to alleviate environmental and food safety concerns over transgenic plants in vegetatively and sexually propagated plants, to prevent volunteer transgenic plants, to protect proprietary transgenic technologies, and to allow farmers to reuse their harvested seeds for future planting.

Indication of Genetic Linkage Map for Sunflower(Helianthus annuus L.) by SSR Markers

In order to investigate the possibility and efficiency of exogenous gene spread in nature and potential ecological risk of transgenic rice, as well as analyze the effect of exogenous Bt gene insertion on ecological fitness of transgenic rice plants, a experiment was carried out with three insect-resistant Bt transgenic rice cultivars Bt63, R1 and R2 and one conventional rice line 11-838 as experimental materials, the insect-resistant transgenic and non-transgenic rice plants were inter- cropped pair-wisely under high and low insect-infestation pressures, and the vegeta- tive growth, seed-setting and the resistance to rice stem borers were compared be- tween transgenic and non-transgenic lines. According to the experimental results, both the tiller number and fresh weight of Bt transgenic rice plants under low insect- infestation pressure showed no significant differences compared with the control, but the plant height, spike length and spike weight were all lower than those of non- transgenic rice plant, and Bt63 and R2 were significantly different compared with the control. On the contrary, under high insect-infestation pressure, the tiller number, spike length and spike weight of three Bt transgenic rice cultivars were significantly higher than those of the control, while the plant height showed different fitness ef- fects among various transgenic rice cultivars, which might be related to the charac- teristics of the receptive cultivars. The individual filled grain number and 1 000-grain weight of three transgenic rice cultivars showed no significant difference compared with the control under two different insect-infestation pressures, suggesting that the effect of exogenous Bt gene on seed setting was not significant. Under insect-infes- tation pressure, the resistance of three Bt transgenic rice cultivars against rice stem borer was significantly superior to non-transgenic rice, indicating that the effect of exogenous Bt gene on insect resistance of receptive plants was distinctly. Further- more, experimental results showed that the fitness cost of Bt transgenic rice was rel- atively low, which implied that exogenous Bt gene in insect-resistant transgenic rice might escape under certain environmental conditions, but this risk was very low.

The antibiotic resistome： gene flow in environments, animals and human beings

The antibiotic resistance is natural in bacteria and predates the human use of antibiotics. Numerous antibiotic resistance genes （ARGs） have been discovered to confer resistance to a wide range of antibiotics. The ARGs in natural environments are highly integrated and tightly regulated in specific bacterial metabolic networks. However, the antibiotic selection pressure conferred by the use of antibiotics in both human medicine and agriculture practice leads to a significant increase of antibiotic resistance and a steady accumulation of ARGs in bacteria. In this review, we summarized, with an emphasis on an ecological point of view, the important research progress regarding the collective ARGs （antibiotic resistome） in bacterial communities of natural environments, human and animals, i.e., in the one health settings. We propose that the resistance gene flow in nature is ＂from the natural environments＂ and ＂to the natural environments＂; human and animals, as intermediate recipients and disseminators, contribute greatly to such a resistance gene ＂circulation.＂

Development and validation of atmospheric gene flow model for assessing environmental risks from transgenic corn crops

Pollen-mediated gene flow from genetically modified plants to non-target plants is a concern of crop growers,seed companies,the general public,and the scientific communities.Although there have been descriptive and mechanistic models to describe pollen dispersion,there has rarely been a comprehensive mechanistic model to dynamically simulate pollen release,dispersion,and deposition and to finally relate them to the gene flow(outcrossing).This research developed and validated such a comprehensive mechanistic model for corn crop gene flow risk management.Dynamic pollen dispersion and deposition were predicted by a 3-D random walk model according to inputs of weather data and plant and domain characteristics.Actual gene flow(outcrossing ratio)was obtained according to predicted grand total deposition flux at silk height during the whole pollination season.The model was validated by experimental data and was appropriate to predict gene flow with acceptable accuracy under different atmospheric and environmental conditions;on average,the ratios of measured and simulated values ranged from 0.82 to 1.21,while R2 ranged from 0.56 to 0.68.The model can be easily adapted for other genetically modified crops.

Genetic boundary and gene flow between 2 parapatric subspecies of brown rats

Two parapatric Rattus norvegicus subspecies,R.n.humiliatus(RNH)and R.n.caraco(RNC),are classified accordi ng to morphological diverge nee and are mainly distributed in North and Northeast China.Here,we aimed to explore the population genetic structure,genetic boundary,and gene flow in these rats using 16 microsatellite loci.Structure analysis and principal component an alysis revealed 3 ancestral clusters.We found that the in termediate cluster exhibited higher genetic diversity and a lower in breeding coefficie nt tha n the other 2 clusters.The genetic differentiation between the 3 clusters was significant but weak,with a higher FST value being observed betwee n the clusters on both sides.The subspecies bou ndary inf erred from microsatellite markers may indicate the existence of an admixture or hybridization area covering Liaoning,Inner Mongolia,and Jilin Provinces,rather than corresponding to the administrative provincial boundaries between Liaoning and Jilin.The RNH and RNC subspecies presented moderate gene exchange and an asymmetric bidirectional gene flow pattern,with higher gene flow from the RNH subspecies to the RNC subspecies,constraining speciation.Such genetic characteristics might be explained by biological processes such as dispersal ability,mate choice,and dynamic lineage boundaries.

Phylogeography of Haplocarpha rueppelii(Asteraceae) suggests a potential geographic barrier for plant dispersal and gene flow in East Africa

East Africa is a biodiversity hotspot. Haplocarpha rueppelii （Sch.Bip.） Beauverd is mainly distributed in the alpine grassland of East Africa. Here we sampled 65 individuals of eight populations/locations of H. rueppelii including hairy and glabrous forms from Mts. Elgon, Aberdare, Kenya, Kilimanjaro and Bale Mountains. We then sequenced one nuclear and three chloroplast DNA fragments and conducted phylogeographic analyses to test the taxonomic rank of the two forms and causes for the differentiation （intrinsic reproductive isolation and geographic barrier）. The results demonstrate that the species consists of two major groups, one includes the populations from Mts. Elgon, Aberdare and Bale, while the other includes Mts. Kenya and Kilimanjaro. The species has established in Mts. Kenya and Aberdare during the Pleistocene. However, migration rate for individuals between the two mountains was low as showed by gene flow analysis. A barrier for plant dispersal and gene flow would have existed between Mts. Aberdare and Kenya since at least Pleistocene. No change of the taxonomic concept of this species is needed. This study reveals a potential geographic barrier in East Africa. We hope it will arouse more scientists＇ interests in phylogeography and biodiversity of East Africa.

Application of an atmospheric gene flow model for assessing environmental risks from transgenic corn crops

Gene flow data from experiments under limited environmental conditions(e.g.wind speed and direction,atmospheric stability)have only provided limited information for gene flow risk management.It is necessary to apply models to predict the gene flow under a complete set of possible environmental conditions to inform farmers,seed companies,government agencies,and researchers about the risks and potential prevention and precaution methods.In this paper,the previous validated gene flow model developed by the authors was used to predict gene flow from genetically modified(GM)corn crops.The model was used to simulate potential gene flow from GM corn sources of different sizes from one plant area of 0.1 m^(2) to an area 3.1×10^(6) m^(2) under normal weather conditions.In addition,the model was also used to predict the potential gene flow for different source strengths,atmospheric conditions,buffer heights,buffer field sizes,and pollen settling speeds from 10,000 m^(2) sources.The model simulations have provided gene flow information for risk management under the above conditions and have shown that the source sizes,source strengths,buffer heights,buffer sizes,atmospheric conditions,and pollen settling speeds had important effects on gene flow.While the atmospheric conditions and pollen settling speeds cannot be controlled,choosing appropriate buffer heights and sizes will effectively prevent gene flow.The lost seed control is crucial to limit gene flow because even a GM corn plant can result in a grand total deposition flux of 646,272 grains/m^(2),an outcrossing ratio of 0.016,and outcrossed seed of 110 kernels/m^(2) at 0.8 m from the plant in the non-target field under normal atmospheric conditions.