Genome-wide recombination variation in biparental segregating and reciprocal backcross populations provides information for introgression breeding in Brassica napus

Variation in patterns of recombination in plant genomes provides information about species evolution,genetic diversity and crop improvement. We investigated meiotic crossovers generated in biparental segregating and reciprocal backcross populations of the allopolyploid genome of rapeseed(Brassica napus)(AACC, 2n = 38). A structured set of 1445 intercrossed lines was derived from two homozygous de novo genome-assembled parents that represented the major genetic clusters of semi-winter Chinese and winter European rapeseeds, and was used to increase QTL resolution and achieve genomic reciprocal introgression. A high-density genetic map constructed with 6161 genetic bins and anchored centromere regions was used to establish the pattern of recombination variation in each chromosome. Around 93%of the genome contained crossovers at a mean rate of 3.8 c M Mb^(-1), with the remaining 7% attributed to centromeres or low marker density. Recombination hotspots predominated in the A genome, including two-thirds of those associated with breeding introgression from B. rapa. Genetic background might affect recombination variation. Introgression of genetic diversity from European winter to Chinese semi-winter rapeseed showed an increase in crossover rate under the semi-winter environment. Evidence for an elevated recombination rate having historically contributed to selective trait improvement includes accumulation of favorable alleles for seed oil content on hotspots of chromosome A10. Conversely, strong artificial selection may affect recombination rate variation, as appears to be the case with a coldspot resulting from strong selection for glucosinolate alleles on A09. But the cold region would be promptly reactivated by crossing design indicated by the pedigree analysis. Knowledge of recombination hotspots and coldspots associated with QTL for 22 traits can guide selection strategies for introgression breeding between the two gene pools. These results and rich genomic resources broaden our understanding of recombination behavior in allopolyploids and may advance rapeseed genetic improvement.

Improvement for Agronomic Traits of Partial Waxy Wheat by Combination of Backcrossing with a PCR-based DNA Marker

To improve agronomic traits of partial waxy wheat, crossing between Chinese Baihuomai and wheat cultivars PH85-16, Jinan 17, and Yannong 15 was performed. The progeny plants were further backcrossed to these cultivars as recurrent parents for five generations. To get homozygous plants with the null allele at the Wx-D1 locus, self-pollination was carried out in the BC5F1 generation. Through another three generations, 6 partial waxy wheat lines were obtained, which had similar agronomic performance as their recurrent parents and carried the null allele at the Wx-D1 locus. In each generation, the Wx-D1 locus was identified by a PCR-based DNA marker and the agronomic traits were examined in progeny plants. The results from this study indicate that the use of backcrossing with a PCR-based DNA marker was useful in waxy wheat breeding. These partial waxy wheat lines can be used in field production.

Identification for Heat Tolerance in Backcross Recombinant Lines and Screening of Backcross Introgression Lines with Heat Tolerance at Milky Stage in Rice

The present study aimed at breeding new rice germplasms with similar genome but significantly differed in heat tolerance during the grain filling stage.A total of 791 BC1F8 backcross recombinant lines,derived from the cross of Xieqingzao B /N22 //Xieqingzao B,were used as materials.Each rice line was separated evenly into two groups,and the heat tolerance of all rice lines were evaluated at natural high temperature in fields.The rice lines with heat tolerant index higher than 90% or lower than 40% were selected to compare the phenotypic characters and further identify heat tolerance at the early milky stage in a phytotron.Rice lines with similar phenotypic characters but significantly differed in heat tolerance at the milky stage were analyzed by 887 simple sequence repeat markers that were evenly distributed on the 12 rice chromosomes.In the result,12(6 pairs) rice lines with similar phenotypic characters but significantly differed in heat tolerance at the milky stage were obtained.Molecular marker analysis indicated that the genomic polymorphism between 703T and 704S was the smallest in the 6 pairs of rice lines,with only 16 polymorphic sites,including 22 different alleles.The application of these two backcross introgression rice lines for future study on the mechanisms of heat tolerance in rice at the milky stage will be theoretically beneficial in reducing the interference caused by genetic differences from experimental materials.

Development of New Submergence Tolerant Rice Variety for Bangladesh Using Marker-Assisted Backcrossing

Submergence tolerant high yielding rice variety was developed using BR11 as a recipient parent applying foreground, phenotypic and background selection approaches. Recombinant selection was found essential to minimize linkage drag by BC2F2 generation. Without recombinant selection, the introgression size in the backcross recombinant lines （BRLs） was approximately 15 Mb on the carrier chromosome. The BRLs were found submergence tolerance compared to the check varieties under complete submergence for two weeks at Bangladesh Rice Research Institute, and produced higher yield compared to the isogenic Subl-line under controlled submerged condition. The BRL IR85260-66-654-Gaz2 was released as BRRI dhan52 in 2010, which was the first high yielding submergence tolerant variety in Bangladesh. BRRI dhan52 produced grain yield ranging from 4.2 to 5.2 t/hm2 under different flash flood prone areas of Bangladesh in three consecutive seasons. The study demonstrated the efficiency of recombinant selection and better adaptability of the newly released submergence tolerant high yielding variety in flash flood prone different areas of the country with respect to submergence tolerance and yield potential.

Identification and QTL mapping of Z550, a rice backcrossed inbred line with increased grains per panicle

An elite backcrossed inbred line Z550 with increased grains per panicle was identified from advanced backcrosses between Nipponbare and Xihui 18 by simple sequence repeat(SSR) marker-assisted selection(MAS). Z550 carries 13 substitution segments distributed on chromosomes 1, 6, 7, 8, 9, 10, and 12, with an average substitution length of 1.68 Mb. Compared with the Nipponbare parental line, plant height, panicle length, spikelets per panicle, grains per panicle, and grain weight for Z550 were significantly increased. While the grain width of Z550 was significantly narrower, and the seed setting ratio(81.43%) was significantly lower than that of Nipponbare, it is still sufficient for breeding purposes. Quantitative trait loci(QTLs) mapping for important agronomic traits was conducted with the F_2 population derived from Nipponbare crossed with Z550 using the restricted maximum likelihood(REML) method. A total of 16, including 12 previously unreported QTLs were detected, with contribution rates ranging from 1.46 to 10.49%. Grains per panicle was controlled by 8 QTLs, 5 of which increased number of grains whereas 3 decreased it. qGPP-1, with the largest contribution(10.49%), was estimated to increase grains per panicle by 30.67, while q GPP-9, with the minimum contribution rate(2.47%), had an effect of increasing grains per panicle by 15.79. These results will be useful for further development of single segment substitution lines with major QTLs, and for research of their molecular functions via QTL cloning.

Improvement of three popular Indian groundnut varieties for foliar disease resistance and high oleic acid using SSR markers and SNP array in marker-assisted backcrossing

Foliar fungal diseases(rust and late leaf spot)incur large yield losses,in addition to the deterioration of fodder quality in groundnut worldwide.High oleic acid has emerged as a key market trait in groundnut,as it increases the shelf life of the produce/products in addition to providing health benefits to consumers.Marker-assisted backcrossing(MABC)is the most successful approach to introgressing or pyramiding one or more traits using traitlinked markers.We used MABC to improve three popular Indian cultivars(GJG 9,GG 20,and GJGHPS 1)for foliar disease resistance(FDR)and high oleic acid content.A total of 22 BC3F4 and 30 BC2F4 introgression lines(ILs)for FDR and 46 BC3F4 and 41 BC2F4 ILs for high oleic acid were developed.Recurrent parent genome analysis using the 58 K Axiom_Arachis array identified several lines showing upto 94%of genome recovery among second and third backcross progenies.Phenotyping of these ILs revealed FDR scores comparable to the resistant parent,GPBD 4,and ILs with high(~80%)oleic acid in addition to high genome recovery.These ILs provide further opportunities for pyramiding FDR and high oleic acid in all three genetic backgrounds as well as for conducting multi-location yield trials for further evaluation and release for cultivation in target regions of India.

QTL and genetic analysis controlling fiber quality traits using paternal backcross population in upland cotton

Background:Genetic improvement in fiber quality is one of the main challenges for cotton breeders.Quantitative trait loci(QTL)mapping provides a powerful approach to dissect the molecular mechanism in fiber quality traits.In present study,F14 recombinant inbred line(RIL)population was backcrossed to paternal parent for a paternal backcross(BC/P)population,deriving from one upland cotton hybrid.Three repetitive BC/P field trials and one maternal backcross(BC/M)field trial were performed including both two BC populations and the original RIL population.Results:In total,24 novel QTLs are detected for fiber quality traits and among which 13 QTLs validated previous results.Thirty-five QTLs in BC/P populations explain 5.01%–22.09%of phenotype variation(PV).Among the 35 QTLs,23 QTLs are detected in BC/P population alone.Present study provides novel alleles of male parent for fiber quality traits with positive genetic effects.Particularly,qFS-Chr3–1 explains 22.09%of PV in BC/P population,which increaseds 0.48 cN·tex−1 for fiber strength.A total of 7,2,8,2 and 6 QTLs explain over 10.00%of PV for fiber length,fiber uniformity,fiber strength,fiber elongation and fiber micronaire,respectively.In RIL population,six common QTLs are detected in more than one environment:qFL-Chr1–2,qFS-Chr5–1,qFS-Chr9–1,qFS-Chr21–1,qFM-Chr9–1 and qFM-Chr9–2.Two common QTLs of qFE-Chr2–2(TMB2386-SWU12343)and qFM-Chr9–1(NAU2873-CGR6771)explain 22.42%and 21.91%of PV.The region between NAU4034 and TMB1296 harbor 30 genes(379 kb)in A05 and 42 genes(49 kb)in D05 for fiber length along the QTL qFL-Chr5–1 in BC/P population,respectively.In addition,a total of 142 and 46 epistatic QTLs and QTL×environments(E-QTLs and QQEs)are identified in recombinant inbred lines in paternal backcross(RIL-P)and paternal backcross(BC/P)populations,respectively.Conclusions:The present studies provide informative basis for improving cotton fiber quality in different populations.

Transfer of Lysozyme Gene into indica Parents of Hybrid Rice by Backcrossing

A lysozyme gene resistant to rice blast was transferred from the donor transgenic japonica rice Zhonghua 9 （D2-1-2） into a sterile line Pei＇ai 64S（PA 64S） and restorer line 9311 of the two-line hybrid rice Liangyoupeijiu, and the restorer line Minghui 63 （MH 63） of three-line hybrid rice Shanyou 63 by successive backcrossing. The PCR analysis confirmed that foreign lysozyme gene was segregated at ratio of 1 ： 1 in backcross generations of B39311, B3MH63 and B2PA64S, and at ratio of 3 ： 1 in selfed generations of B2F2 9311, B2F2 MH63 and B1F2 PA64S, indicating that the foreign gene was stably inherited over successive generations as a dominant single copy gene. The resistance against rice blast in backcross or selfed generations and corresponding testcross combinations were investigated in 2003 and 2004. The results showed that the resistance of the transgenic rice to blast had a greater improvement than that of the corresponding recurrent parents or the corresponding check hybrid combinations. The resistance of the advanced backcross and selfed generations to rice blast is much stronger than that of the early generations. The study confirmed that transferring the lysozyme gene into hybrid parents by backcrossing was a simple and effective approach to develop new hybrid rice resistant to rice blast.

Application of Marker Assisted Backcrossing to Introgress the Submergence Tolerance QTL <i>SUB</i>1 into the Vietnam Elite Rice Variety-AS996

The result of the study contributes to enhancing and sustaining future livelihoods and food security in Vietnam vis-a-vis climate change. An innovative strategy based on marker-assisted backcrossing (MABC) was used to transform popular rice variety AS996 into the one can tolerate submergence while maintaining its original characteristics preferred by farmers and consumers. The submergence tolerance QTL SUB1 counts for up to 70% of the submergence tolerant and provides a marked improvement of submergence tolerance in all genetic backgrounds and environments tested so far. Parental diversity was carried out with 460 markers. Of which, 53 polymorphic markers were used for assessment on BC1F1, BC2F1 and BC3F1 generations. The best BC1F1 plant was P422 with highest recipient allele was 87.5%, while the additional chosen plants were P412, P428, P215 and P39 (81% - 84%). All these plants were used to develop BC2F1 generation. The six BC2F1 plants were used to develop BC3F1 and BC2F2 were the plants number P422-11 and P422-14 having 93.75% recipient alleles and P422-12, P422-3, P39-17, P39-25 having 92.25% recipient alleles. Total of 445 BC3F1 plants were confirmed the introgresion of SUB1 using ART5 and SC3. After three generations of backcrossing, application of MABC resulted in the best BC3F1 individual P422-14-177 with 100% of recipient alleles based on the number of 53 markers used with only the introgression size of SUB1 was 0.3Mb between ART5 and SC3. Phenotyping was carried out on BC3F1 and BC2F2 of the selected lines. The survival ratio of these selected lines and IR64SUB1 were the same. It convinced the successfully introgress SUB1 into AS996 rice variety. The breeding line BC4F1 having 100% genetic background of donor variety is ready for develop new submergence tolerant rice variety ASS996-SUB1 to cope with climate change.

The genetic variation of the backcross modified lines developed from the maize line 08-641 selected by different directions

In this study, two donaors CN962 and 8065 were used to improve the resistance to northern leaf blight of the recurrent parent inbred line 08-641 (R08). A total of 79 lines (BC2F4) were developed by a bidirectional selection based on the similarity and dissimilarity in the shape and color of seeds to R08.The genetic variation of these lines were analyzed by 44 pairs of SSR molecular markers, the result showed that a total of 272 alleles were detected in the improved lines and R08, 123 out of them were detected in the modified lines but discarded in R08. The modified line selected based on dissimialry in the shape and corlor of seeds to R08 have lower genetic similary between R08 than that between the lines selected based on similary in the shape and color of seeds and R08, and the genetic variation of these lines were wider. It concluded that when the backcross breeding were used to improve the maize inbred lines, multidirectional selection based on phenotypic value were contribute to create and keep genetic variation.

Combined Selection in Backcross Population of Papaya(Carica papaya L.)by the Mixed Model Methodology

The selection of superior genotypes based on the simultaneous response to several characteristics of agronomic importance is a key strategy to overcome the scarcity of available varieties of papaya. This study aimed to apply the combined selection by using distinct selection indexes based on both the genetic values obtained by the REML/BLUP methodology and the real measured values to select agronomically superior genotypes of papaya within backcross progenies. The combined selection was carried out based on genetic and phenotypic values, original and standardized, multiplied by the agronomic weights. The results of the analysis of genetic parameters indicate that the evaluated progenies have expressive genetic variability for the considered traits, and that there are real possibilities of genetic progress with the selection. Among the analyzed indexes, the one based on standardized genetic value presented greater consistency in the ranking of genetic material, demonstrating the advantage of data standardization. Five progenies belonging to the BC1 generation, and five to the BC3 generation were selected using this index. A total of 27 plants ag-ronomically superior were selected within the top five progenies and recommended for generation advance, 23 being selected by combined selection and 4 using the direct selection for the four mainly characters in papaya breeding program: production, pulp and fruit firmness and soluble solids. Beyond the selection of superior genotypes for the development of future inbred lines, this study also allowed defining the best strategy to apply the combined selection in papaya using pre-dicted breeding values obtained by BLUP. This strategy may allow higher accuracy in the selection process, thus increasing the chances of success of the breeding programs.

Core Collection Based Backcrossing: An Eff icient Approach for Breeding, Germplasm Enhacement and Gene Discovery

Plant germplasm underpins much of crop development. Millions of germplasm accessions have been collected and conserved ex situ, and the major challenge is now how to exploit and utilize this abundant resource.

甘蓝型油菜pol CMS恢复系621R硼高效利用的遗传改良

油菜是需硼量最大的作物之一,在生产中需要额外增施硼肥,否则易出现大幅度减产。为了提高油菜对硼元素的利用效率,本研究以携带Bna.A03NIP5;1优良等位基因的硼高效利用材料2016-XFSB为供体亲本,以621R为轮回亲本,通过回交与MAS相结合的方法,将Bna.A03NIP5;1优良等位基因导入621R中,选育出拥有621R遗传背景并携带有Bna.A03NIP5;1优良等位基因的近等基因系材料621R^(NIP5;1A3),并在此基础上与细胞核雄性不育系RG430A配制了杂交种。利用水培和不同土质土壤栽培试验对621R和621R^(NIP5;1A3)及其杂种进行相关农艺性状的鉴定。结果表明,621R^(NIP5;1A3)及其杂交种对硼的利用效率有明显提升。农艺性状考察结果表明,621R^(NIP5;1A3)及其改良杂交种在生物量、株高、角果长、每角果粒数和千粒重等性状上有显著优势。本研究结果对油菜硼高效育种具有重要参考价值。

Characterization of interspecific hybrids and backcross progenies from a cross between Oryza minuta and Oryza sativa

Oryza minuta, a tetraploid wild relative of cultivated rice, is an important source for the genetic improvement. Interspecific hybrids were obtained from the cross of O. sativa L. (IR24) and O. minuta (Acc. No. 101133) with 5.58% crossability, which ranged from 0.11% to 1.62% in the backcross generations. The chromosome numbers of the backcross progenies were 24 to 48. Seven yield-related traits of the parents, hybrid F1, and backcross progenies were evaluated. Simple sequence repeat markers analysis showed that the polymorphism ratio of SSR bands between IR24 and Acc. No. 101133 was 93.2%. The average donor segment number, length, donor genome size, and percentage of donor genome of 92 BC3F1 plants (2n=24) were 24.1, 17.8 cM, 438.4 cM and 26.2%, respectively. They were complex variation and uneven among the chromosomes. These introgression lines could be used to identify the favorable genes of O. minuta and provide a new platform for the genetic improvement of cultivated rice.

不同遗传背景的转Cry1C基因玉米品系的室内抗虫性鉴定

以转Cry1C基因的转基因玉米C492为供体,通过回交转育的方式将目标基因分别导入不同遗传背景的玉米自交系,获得转基因玉米自交系C492/N-76、C492/N202、C492/S273、C492/DH1901和C492/DH1904,采用室内离体组织生测的方法研究各品系对玉米螟和草地贪夜蛾的抗虫性。结果表明:转基因玉米C492和5个不同遗传背景自交系均表现出高抗玉米螟,虽对草地贪夜蛾的抗性有一定差异,但接虫6 d后幼虫死亡率均达到80%以上。这说明转基因玉米C492和5个不同遗传背景自交系可用于抗虫转基因玉米新品种培育,具有较好的商业化应用前景。

Creating Wheat Germplasm for High Quality Breeding by Monosomic Backcrossing

By crossing bread wheat cultlvar GC8901 with the 1D monosonlc line of Xiaoyan No. 6 and backcrosslng the offsprlng with the Xlaoyan No. 6 1D monosonlc llne for 5 years, high-molecular-welght glutenin subunlts 1Dx5＋1Dy10 from GC8901 have been transferred Into wheat cultivar Xiaoyan No. 6. The BC5F1 offspring lines had been detected by using methods of cytology, marker, molecular marker and six elite single plants with high molecular-welght glutenin subunlts： lAx1, 1Bx14＋1 By15, 1Dx5＋1 Dy10 were Identified. Those lines have high-yleld potential with better agronomic characters and have been used In high quality wheat breeding processes as well.

Statistical method for mapping QTLs for complex traits based on two backcross populations

Most important agronomic and quality traits of crops are quantitative in nature.The genetic variations in such traits are usually controlled by sets of genes called quantitative trait loci (QTLs),and the interactions between QTLs and the environment.It is crucial to understand the genetic architecture of complex traits to design efficient strategies for plant breeding.In the present study,a new experimental design and the corresponding statistical method are presented for QTL mapping.The proposed mapping population is composed of double backcross populations derived from backcrossing both homozygous parents to DH (double haploid) or RI (recombinant inbreeding) lines separately.Such an immortal mapping population allows for across-environment replications,and can be used to estimate dominance effects,epistatic effects,and QTL-environment interactions,remedying the drawbacks of a single backcross population.In this method,the mixed linear model approach is used to estimate the positions of QTLs and their various effects including the QTL additive,dominance,and epistatic effects,and QTL-environment interaction effects (QE).Monte Carlo simulations were conducted to investigate the performance of the proposed method and to assess the accuracy and efficiency of its estimations.The results showed that the proposed method could estimate the positions and the genetic effects of QTLs with high efficiency.

回交改良中粳616抗除草剂性状

除草剂的使用会降低水稻的产量和品质,开展非转基因的抗除草剂水稻品种改良迫在眉睫。以中粳616(受体材料)及抗除草剂品种金粳818、净田美占为试验材料,经除草剂(先达豆施乐,5%咪唑乙烟酸含量)浓度梯度试验筛选抗除草剂处理浓度,通过杂交、回交、自交结合除草剂耐药性筛选获得中粳616-TR,进一步分析除草剂对改良后中粳616-TR幼苗萌发及生长的影响,同时以中粳616为对照进行品比试验。结果表明:除草剂浓度梯度试验结果显示,可将3 mL·L^(-1)作为后续抗除草剂水稻筛选浓度。改良获得的中粳616-TR在3 mL·L^(-1)浓度除草剂处理下,在萌发期和苗期对除草剂均有耐药性,品比试验发现中粳616-TR在农艺性状、产量、抗病性、品质方面与中粳616无显著差异。综上,改良后的中粳616-TR具有品质优、稻瘟病抗性好、产量高兼备抗除草剂性状等特点。

基于KASP平台的转基因玉米高通量特异性检测方法

为了促进转基因玉米产业化发展,加快优良性状转化体与骨干自交系的转育工作,适用于回交群体的高通量前景选择方法亟待开发。本研究以转基因玉米DBN9936为材料,根据其外源插入片段及其侧翼序列,采用primer5软件设计了6对特异性引物,与内标准基因zSSIIb引物进行组合用于评估,获得最优引物组合后,进一步开展特异性验证、检出限测试和多样品验证。结果显示,最优引物组合为DBN9936-LB1*zSSIIb-k1;10份DBN9936 BC1代种子的KASP基因分型结果与琼脂糖凝胶电泳结果完全一致;10份转化体DBN9858、DBN9501、C0010.1.3、C0010.3.1、C0010.3.7、C0030.2.5、BT11、GA21、MIR162和2A-7的转基因玉米样品在双平台上检测结果均表现为阴性;转基因玉米DBN9936的检出限为10%;48份不同DBN9936杂交种材料的KASP基因分型结果均表现为阳性。综上,本研究方法可用于转基因玉米DBN9936回交转育过程中的前景选择,且样品通量高,为其他农作物在转基因育种过程中的目标基因检测提供了技术参考。

‘滁菊’回交BC1株系的遗传变异与综合评价

[目的]本文旨在通过回交创制优质稳产的茶用菊新种质,并对获得的BC1群体进行遗传分析和评价,以筛选综合性状优良或特定性状突出的优异材料应用于进一步的育种改良。[方法]将‘苏菊14号’(‘滁菊’杂交1代)与‘滁菊’回交获得BC1株系群体。对102个BC1扩繁株系的12个相关产量、外观性状以及绿原酸、木犀草苷、3,5-O-双咖啡酰基奎宁酸和总黄酮4种功能性成分含量进行测定和分析,再运用灰色关联度分析法对BC1群体进行综合评价。[结果]大多数BC1群体花朵分布特征和田间适应性表现优异且遗传稳定。着花部位为偏母性遗传,花期茎枝直立程度、田间生长势、花期3个性状为偏父性遗传。12个性状的灰色关联分析权重值以田间适应性、花朵分布特征的权重值最大。利用评价结果结合聚类分析将102个回交BC1株系分为优、良、中、差4个级别。BC1群体中4种功能性成分含量均达到药用菊花规定的株系有8个,占比为7.84%,可作为茶、药兼用种质。通过综合评价,共筛选出4个优良株系,其中BC1-97的功能性成分含量水平优于‘滁菊’,其余3个株系接近‘滁菊’的功能性成分含量水平。[结论]本研究通过遗传分析、灰色关联度分析和功能性成分测定,在对‘苏菊14号’与‘滁菊’回交BC1群体综合评价的基础上,共筛选出4个优良株系,为‘滁菊’的进一步回交改良提供了中间种质和理论依据。