Correlation between Stigma Characteristics and Outcrossing Rate of Five Photo-thermo Sensitive Genic Male Sterile(PTGMS) Rice Lines

[Objective] This study aimed to investigate the correlation between stigma characteristics and outcrossing rate of photo-thermo sensitive genic male sterile（PTGMS） rice lines. [Method] The stigma exsertion rate, stigma size and stigma vitality of Chuang 5S, Yun 9S, Peiai 64 S, P88 S and Y58 S were analyzed. Using R608 as a male parent, outcrossing rate of these five PTGMS lines was determined. [Result]There were differences in stigma exsertion rate, stigma size and stigma vitality among five PTGMS lines. P88 S exhibited the best stigma characteristics and relatively strong stigma vitality, with the stigma exsertion rate of 95.94%, stigma vitality coefficient of 2.71, outcrossing rate of 84.02% in the field. Peiai 64 s had the minimum stigma length（2.41 mm） and stigma width（1.10 mm） and the weakest stigma vitality, with the stigma exsertion rate of 93.04%, outcrossing rate of 65.47%. Yun 9S,Y58 S and Chuang 5S displayed medium stigma characteristics, among which Yun9 S exhibited higher outcrossing rate than other two PTGMS lines. Correlation analysis showed that stigma exsertion rate, stigma length, stigma width, stigma area and stigma volume were significantly correlated to outcrossing rate（P〈0.05）, with correlation coefficients of 0.897, 0.934, 0.934, 0.939 and 0.937, respectively. However,there was no significant correlation between stigma vitality coefficient and outcrossing rate. [Conclusion] Based on stigma characteristics of five PTGMS lines, auxiliary pollination measures should be adopted reasonably to save production costs and improve hybrid rice seed yield.

Breeding and Utilization of Japonicalinous CMS Line Chunjiang 19A with Wide Compatibility and High Outcrossing Rate

[Objective] To breed and utilize the japonicalinous cytoplasmic male sterile （CMS） line Chunjiang 19A with wide compatibility and high outcrossing rate. [Method] Cbunjiang 16A was used as the female parent to cross with Indica-Japonica crossing material B125, which was used as male parent. Backcross breeding was contin- ued for consecutive generations to breed the CMS lines with wide compatibility and high outcrossing rate. [Result] Chunjiang 19A is a late medium Japonica CMS line with good fertility stability. The proportion of sterile plants and pollen abortion rates are 100% and the Cheng＇s index is 14. It is the japonicalinous CMS line with early flowering, partial stigma exsertion, high outcrossing rate, wide compatibility, high re- sistance to stripe disease and good combining ability. The hybrid rice combinations originated from Chunjiang 19A have strong tUlering ability, dominant heterosis, high resistance to stripe disease, great yield potential and good color change at mature stage. [Conclusion] The successful breeding of Chunjiang 19A laid good foundation for the breeding of Japonica hybrid rice and Indica-Japonica hybrid rice combinations with high seed production and dominant heterosis, showing a promising application prospect.

Effect of High Temperature on Outcrossing Characteristics at the Fertility Sensitive Period of Photo-thermo Sensitive Genic Male Sterile (PTGMS) Rice Lines

Four sterile lines （Peiai64S, Y58S, Guangzhan 63-2S and H638S） and the restoring line R527 were used as materials. Five temperature gradients （24, 27, 30, 33 and 36 ℃ in artificial climate chamber） and the natural temperature （as control） were treated to the four sterile lines for 6 d in the fertility sensitive period of heading to flowering stage, respectively, to study the effects of temperature on physiological biochemical characteristics of young panicles and outcrossing characteristics. The results showed that the percentages of exerted stigma of Peiai 64S and Y58S were the highest at 27 ℃, which were 86.81% and 86.06%, respectively, while the percentages of exerted stigma of Guangzhan 63-2S and H638S were the highest at 24 ℃, which were 76.24% and 81.76%, respectively; the stigma viability of Peiai 64S, Y58S, Guangzhan 63-2S and H638S were the best at 24 ℃, which were 1.96, 2.12, 1.74 and 1.94, respectively; the outcrossing rates of Peiai 64S, Guangzhan 63-2S and H638S were the highest at 24 ℃, which were 58.87%, 54.22% and 50.50%, respectively, while the outcrossing rate of Y58S was the highest at 27 ℃, which was 58.96%; and the contents of peroxidase （POD） of the four sterile lines increased significantly at 33 ℃ compared with the control, while the contents of malondialdehyde （MDA） and proline increased significantly at 36 ℃ compared with the control. There were differences in temperature sensitivity between the male sterile lines, and the 24 ℃ treatment during the sensitive period was the best for the fertility sensitive period of Peiai 64S, while 27 ℃ was the best temperature for Y58S, Gangzhan 63-2S and H638S.

A microsatellite study on outcrossing rates and contamination in an Eucalyptus globulus breeding arboretum

Four pairs of microsatellite molecular polymorphism primers were used to analyse microsatellite fingerprints of 188 seedlings derived from an open-pollinated progeny grafted Eucalyptus globulus breeding arboretum in Victoria, south-eastern Australia. The microsatellite loci chosen for this study were highly polymorphic with the mean number of alleles per locus of 14.25. Individual mothers varied in their outcrosssing rate estimate from 15% to 95%, the overall outcrossing level in the arboretum was 47.9% and the contamination rate was 17.6%. The high selfing level was likely to result in marked inbreeding depression in the performance of open-pollinated seed lots. Open-pollinated seeds collected from such arboreta are not advisable because of its low genetic quality, although such arboreta may be useful for the seed production through large-scale manual pollination or collecting seeds only from trees or genotypes within the arboretum that have high outcrossing rates.

Genetic effects and plant architecture influences on outcrossing rate in soybean

Outcrossing rate is an important determinant of cytoplasmic male sterile(CMS)breeding and hybrid seed production for heterosis in soybean.Parental lines with a high outcrossing rate were screened for backcross breeding to obtain the high outcrossing rate maintenance B-lines and sterile A-lines.Application in production practices will help to increase hybrid soybean production.In this study,JLCMS82B and JLCMS89B were selected as parents for the construction of outcrossing rate segregation populations,and the progeny-array approach(PAA)and glyphosate resistant gene markers were used to determine outcrossing rates.We found that:(1)The outcrossing rate between JLCMS82B and JLCMS89B was significantly different;(2)the outcrossing rate of the F2 segregating populations was a quantitative trait,though whether an additive or epistatic effect exists required analysis with a triple test intersection analysis;(3)agronomic traits correlated with outcrossing rate;outcrossing rate was the highest with plant height of about 84 cm,lower number of plant branches,earlier flowering time,larger angle between the branches and the main stem,and with more divergent plant morphology.Correlation analysis between agronomic traits and outcrossing rate can effectively guide the screening of parents with a high outcrossing rate.

New Temperature Sensitive Genic Male Sterile Lines with Better Outcrossing Ability for Production of Two-Line Hybrid Rice

An investigation was carried out with three newly developed temperature sensitive genic male sterile （TGMS） lines for their floral traits, seed production potential and outcrossing ability in ten cross combinations. In the TGMS lines, fertile pollens had an average diameter of 0.89 mm while the sterile pollens was with 0.02 mm diameter.TS-29-150GY produced the biggest fertile pollens with 0.92 mm and other two lines produced relatively smaller pollens with 0.91 and 0.85 ram. Pollen fertility during the fertility reversion period was an average of 60.7%. TS-29-150GY had the maximum of 66.9% spikelet fertility whereas other two lines （TNAU18S and TNAU60S） had relatively lower spikelet fertility of 27.8% and 26.7%, respectively. Average of 17.00 g of seed yield was obtained in the TGMS lines during the fertility reversion period. TS-29- 150GY had the highest value of 21.20 g of seed yield while TNAU18S and TNAU60S produced 16.6 g and 13.2 g of seed yield, respectively. The low seed production ability of these three TGMS lines was attributed only to the environmental conditions prevailing during the period. All three TGMS lines had considerable outcrossing potential of 41.2%, 24.6% and 25.0%, respectively. The cross combinations viz. TNAU18S/IET21508 （36 g/plant）, TNAU18S/IET21044 （13 g/plant）, TNAU18S/IET21009 （26.5 g/plant）, TNAU60S/CB-09-106 （26.2 g/plant）, TNAU60S/IET21009 （14 g/plant） and TS29-150-GY/DRR 3306 （39.2 g/plant） showed perfect synchronization with acceptable hybrid seed yield, indicating suitability of TGMS system under Indian condition. Based on the outcrossing related traits viz. panicle exertion, angle of glume opening, stigma length and pollen size, TNAU18S was identified as the best, followed by TS-29-150GY.

Genome-wide identification of agronomically important genes in outcrossing crops using OutcrossSeq

Many important crops(e.g.,tuber,root,and tree crops)are cross-pollinating.For these crops,no inbred lines are available for genetic study and breeding because they are self-incompatible,clonally propagated,or have a long generation time,making the identification of agronomically important genes difficult,particularly in crops with a complex autopolyploid genome.In this study,we developed a method,OutcrossSeq,for mapping agronomically important loci in outcrossing crops based on whole-genome low-coverage resequencing of a large genetic population,and designed three computation algorithms in OutcrossSeq for different types of outcrossing populations.We applied OutcrossSeq to a tuberous root crop(sweet potato,autopolyploid),a tree crop(walnut tree,highly heterozygous diploid),and hybrid crops(double-cross populations)to generate high-density genotype maps for the outcrossing populations,which enable precise identification of genomic loci underlying important agronomic traits.Candidate causative genes at these loci were detected based on functional clues.Taken together,our results indicate that OutcrossSeq is a robust and powerful method for identifying agronomically important genes in heterozygous species,including polyploids,in a cost-efficient way.The OutcrossSeq software and its instruction manual are available for downloading at www.xhhuanglab.cn/tool/OutcrossSeq.html.

Family Outcrossing Rates of Pinus caribaea Morelet var. caribaea in Seed Orchard and Natural Populations

Family outcrossing rates of Pinus caribaea var. caribaea populations that differed in origin and management were estimated by using isozyme markers. The family outcrossing rates were estimated by (1)fixing the probability of pollen gene frequency (p); (2) allowing pollen gene frequency to vary among families. The estimates of family outcrossing rates for all the populations varied widely among families (clones of the seed orchard), ranging from 0.39 to 2.0. The average family outcrossing rates of both m...

Floral display and breeding system of Jatropha curcas L.

Plant flowering and breeding characteristics are important for us to understand the reproduction of plant populations. In this paper, we studied the reproduction characteristics of Jatropha curcas in Yuanjiang County （23°36＇1＇4, 101°00＇E）, Yunnan Province. The plant produces flowers in dichasial inflorescences. Normally, the flowers are unisexual, and male and female flowers are produced in the same inflorescence. Only a few male flowers are produced in an inflorescence, and fruits are produced only through pollination between different flowers from the same or different plants. By the treatments of emasculation, bagging and artificial pollination in this experiment, there were few but same fruit set ratios when the inflorescences were emasculated, bagged, or bagged with net, except artificial pollination treatments, which showed that Jatropha curcas could produce fruit through apomixis but not wind pollination. When the inflorescences were unbagged, unemasculated and with free pollination treatments, or bagged, emasculated and with artificial cross-pollination treatments, or unbagged, emasculated and with free pollination treatments, there were many fruits produced. It showed that Jatropha curcas shows outcrossing, is self-compatible, and demanding for pollinators. Normally, the male flowers open first and a few flowers bloom in one day in a raceme. These flowers last a long time in bloom. However, a large number of female flowers open from the third to the fifth day, with some female flowers opening first in a few raceme. This shows a tendency to promote xenogamy and minimize geitonogamy.

Mating system and genetic diversity of a rare desert legume Ammopiptanthus nanus (Leguminosae)

Ammopiptanthus nanus is an endangered evergreen shrub endemic to the deserts of central Asia and plays an important role in delaying further desertification. We examined allozyme variation and AFLP diversity in A. nanus populations and investigated the mating system of this species using progeny arrays assayed for polymorphic allozyme loci. Mating system analysis in the Keyi＇eryongke＇er population showed low levels of outcrossing, and strong inbreeding depression. Low levels of genetic variation were detected at both population （allozyme, Pp=14.0%, A=l.14, He=0.031; AFLP, Pp=14.5%, Shannon＇s information index I=0.063） and species （allozyme, Pp=21.1%, A=1.21, He=0.040; AFLP, Pp=20.9%, I=0.083） levels; while moderate genetic differentiation existed among populations, as indicated by allozymes （Gsa-=0.081） and AFLP （GST=0.151-0.193）. Founder effect, bottlenecks in evolutionary history, the mixed mating system and co-ancestry may have influenced the level of genetic diversity in A. nanus. Markers of both types provide new insights for conservation management, indicating that the Biao＇ertuokuoyi and Keyi＇eryongke＇er populations should be given priority for in situ conservation and regarded as seed sources for ex situ conservation.

Factors Influencing the Outcrossed Seed-Setting Obstacles of Photo-thermo Sensitive Genie Male Sterile Rice

Using photo-thermo sensitive genie male rice (PTGMS) Pei' ai 64S, W7415S, W6154S, N26S, Annong S, Nongken 58S, 7001S and 5088S as female parents and conventional indica lines 8258 and U89 as male parents, the factors affecting outcrossed seed-setting were analyzed. The PTGMS had obstacles in outcrossed seed setting influenced by inheritance and environment at varying degrees. Environmental temperature was regarded as the main factor that resulted in the outcrossed seed-setting obstacles. The sensitive stage was at the early stage of grain filling for outcrossed seed setting. There existed remarkable differences at the sensitivity stage, the duration of sensitive period, the sensitive level and the effective level of outcrossed seed-setting obstacles caused by environmental temperature among different PTGMS lines. Therefore, attention should be paid to outcrossed seed-setting obstacles in selection and utilization of PTGMS lines.

Potential Inbreeding in a Small Population of a Mass Flowering Species, <i>Xanthorrhoea johnsonii</i>(Xanthorrhoeaceae): Is Your Mother My Father?

Xanthorrhoea johnsonii is a long lived slow growing perennial understorey species, that produces a large quantity of passively dispersed seed every 3 - 5 years. Reproductive maturity is not reached until 20 - 30 years of age. The temporal asynchrony of the flowering event in this population was analogous to geographic isolation through fragmentation. A small population of plants flowering in isolation provided the opportunity to examine outcrossing rates, genetic diversity and the paternity of progeny at a small spatial scale (0.2 ha). The geographic location and physical characteristics of the adult plants were recorded, and both adults and their seed were sampled for genetic analysis. Four microsatellite loci were screened for genetic diversity and spatial structure analysis. A population outcrossing rate was estimated, as well as the number of paternal parents required to resolve the progeny multilocus genotypes. High genetic diversity was found in both adults and progeny with an estimated 97% outcrossing rate. All maternal lines required several paternal contributors, with no evidence of dominant paternal genotypes. Pollen transfer occurred between both geographically close and distant plants.

Non-associative learning underlies pollination interaction of pollinators and flowering plants

The behavioral response of pollinators is significantly influenced by the prior experience of flower visiting.Learning of pollinators,including non-associative learning,associative learning,and operant conditioning,is determined by the presence or absence of rewards during the flower visiting experience.Here,we indicate that process of non-rewarding flower(empty flower)visiting coincident well with the behavioral paradigm of non-associative learning.Habituation,one of non-associative learning,most likely modulates the pollinating behavior patterns of empty flower visitation.Moreover,we propose that the process of habituation recovery,including spontaneous recovery and dishabituation,may also modulate the behavior of pollinators,which leads to ecological consequences of long-distance pollen dispersal and high outcross pollination rate.We believe that utilizing the methodology of non-associative learning behavioral neurobiology paradigm to investigate pollinator behavior will establish novel insights into the sensory responses and neural activity of pollination behavior in the pollination systems.

OcBSA:An NGS-based bulk segregant analysis tool for outcross populations

Constructing inbred lines for self-incompatible species and species with long generation times is challenging,making the use of F1 outcross/segregating populations the main strategy for genetic studies of such species.However,there is a lack of dedicated algorithms/tools for rapid quantitative trait locus(QTL)mapping using the F1 populations.To this end,we have designed and developed an algorithm/tool called OcBSA specifically for QTL mapping of F1 populations.OcBSA transforms the four-haplotype inheritance problem from the two heterozygous diploid parents of the F1 population into the two-haplotype inheritance problem common in current genetic studies by removing the two haplotypes from the heterozygous parent that do not contribute to phenotype segregation in the F1 population.Testing of OcBSA on 1800 simulated F1 populations demonstrated its advantages over other currently available tools in terms of sensitivity and accuracy.In addition,the broad applicability of OcBSA was validated by QTL mapping using seven reported F1 populations of apple,pear,peach,citrus,grape,tea,and rice.We also used OcBSA to map the QTL for flower color in a newly constructed F1 population of potato generated in this study.The OcBSA mapping result was verified by the insertion or deletion markers to be consistent with a previously reported locus harboring the ANTHOCYANIN 2 gene,which regulates potato flower color.Taken together,these results highlight the power and broad utility of OcBSA for QTL mapping using F1 populations and thus a great potential for functional gene mining in outcrossing species.For ease of use,we have developed both Windows and Linux versions of OcBSA,which are freely available at:https://gitee.com/Bioinformaticslab/OcBSA.

Effects of distinct pollinators on the mating system and reproductive success in Incarvillea sinensis,an annual with large floral displays

Aims Most flowering plants engage in mutualisms with animals to move pollen between individuals,and it is expected that pollinators play an essential role in the evolution of selfing,yet few studies have deter-mined how distinct pollinator types affect a plant’s mating system and reproductive success differentially.We investigated the effect of two different pollinators on the reproductive success of Incarvillea sinensis,an annual with showy,insect-pollinated,one-day flowers.Methods We marked flowers after a single visit from both pollinator species(Bombus patagiatus and Amegilla quadrifasciata)and calculated the number of pollen grains deposited on the stigma and pollen left in the anthers,the fruit and seed set,the outcrossing rate and the cor-relations of paternity.Important Findings There was no significant difference in the number of pollen grains deposited on the stigma,or in the seed and fruit set between the two pollinators in both years.The mean number of pollen grains removed by B.patagiatus was significantly higher than that removed by A.quadrifasciata.The outcrossing rate of flowers pollinated by B.patagiatus was significantly higher than that of flowers pollinated by A.quadrifasciata.The correlation of paternity of flowers pollinated by B.patagiatus was significantly lower than that of flowers polli-nated by A.quadrifasciata.For I.sinensis,B.patagiatus may exhibit larger pollen wastage,thus reducing the male reproductive success of the plant,but it causes higher female reproductive success because of its higher outcrossing rate and the lower correlation of paternity.Our findings highlighted that different animal pollinators could indeed cause differential mating systems and reproductive success.

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.

Genetic Diversity and Mating System Analysis of Pinus massoniana in Second-Generation Clonal Seed Orchard

A total 61 clone parents and 320 open-pollination progenies from 8 clone individuals were identified by 12 polymorphic SSR loci.The result showed that the level of genetic diversity in progeny population was the same as in maternal population, progeny population had all alleles detected in maternal population,and there were not obvious surplus phenomenon of homozygotes in progeny population (F= 0.046).The south crown had more strobili than the north crown,however the genetic diversity of the north crown strobili was not substantially reduced.The progeny of neutral trees had the same level of genetic diversity as in the progeny of partial female trees,with the fixation index tended to be zero,which was accorded with Hardy-Weinberg equilibrium.The multilocus outcrossing rate in the seed orchard was 1.097,and there was no significant inbreeding between parents(t_m-t_s= - 0.031).The multilocus outcrossing rate and singlelocus outcrossing rate in the south crown were higher than that in the north crown.The multilocus outcrossing rate of partial female trees was the same as in neutral trees,and the neutral tree had no obvious declining outcrossing rate with reduced proportion of female and male strobilus compared with partial female tree, with the fixation index tended to be zero.The progeny of the second generation clonal seed orchard still had rich genetic diversity,gene exchange among clones was relatively sufficient,and biparental inbreeding was not significant.