Analysis of genetic and genotype environment interaction effects from embryo, cytoplasm and maternal plant for oleic acid content of Brassica napus L.

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1 _ Plant Science 167 (2004) Analysis of genetic and genotype environment interaction effects from embryo, cytoplasm and maternal plant for oleic acid content of Brassica napus L. Haizhen Zhang a, Chunhai Shi a,, Jianguo Wu a, Yuling Ren a, Changtao Li a, Dongqing Zhang b, Yaofeng Zhang b a Department of Agronomy, College of Agriculture and Biotechnology, Zhejiang University, Hangzhou , China b Crop Research Institute, Zhejiang Academy of Agricultural Sciences, Hangzhou , China Received 29 June 2003; received in revised form 20 December 2003; accepted 26 February 2004 Available online 23 March 2004 Abstract The genetic effects including genetic main and genotype environment interaction effects (GE) for oleic acid content of rapeseed (Brassica napus L.) were studied with eight parents and their F 1 and F 2 during by using genetic model for quantitative traits of seed in diploid plant. The results indicated that the heredity of oleic acid content (OAC) of rapeseed was simultaneously controlled by diploid embryo nuclear genes, cytoplasmic genes and diploid maternal plant nuclear genes as well as their GE interaction effects. Genetic main effects for OAC were more important than GE interaction effects. The total narrow-sense heritability for OAC was high (77.5%), and the general heritabilities were found to be larger than the GE interaction heritabilities. The predicted genetic effects indicated that Huashuang 3 and Zhong R-888 were better than other parents for improving OAC of offspring Elsevier Ireland Ltd. All rights reserved. Keywords: Quality; Oleic acid; Genetic effect; Heritability; Rapeseed (Brassica napus L.) 1. Introduction Oilseed rape, Brassica napus L., is one of the most important oilseed crops around world. Most of the rapeseed oil is produced for human consumption in form of margarine, tried and backed products, and salad oil [1]. The development of oilseeds with altered lipid composition has been the subject of intensive research in recent years due to the industrial and nutritional importance of plant oil. The functional characteristics and quality of the seed oil are primarily determined by the proportion of its main constituent fatty acids and one of the essential components of oilseed fatty acid is oleic acid (C18:1 9) [2]. In order to improve the efficiency of breeding for nutrient quality of rapeseed, understanding the variation for the expression of genes in different environments is necessary. The genetic studies for oleic acid content (OAC) Corresponding author. Tel.: ; fax: address: chhshi@zju.edu.cn (C.H. Shi). has showed that the selection of offspring was efficient because of the high heritability for this quality trait, although the large number of genes were involved in the inheritance of this trait [1,3,4]. The results of Zhou and Liu showed that OAC was mainly controlled by the nuclear genes of the embryo and had partial dominant effect [5]. It was also reported that OAC of rapeseed was dominated by two loci with two alleles per locus [5,6]. The additive effect was the main genetic effect for OAC, and the dominant effect was relatively small [3,4,6]. The high heritability indicated that selection for genotypes with high OAC was efficient. The plants of rapeseed grown under natural environments will be affected by environmental conditions but the phenotypic variation for performance of C18 fatty acids will be affected mainly by genetic effects [7,8]. Shi et al. found that the erucic acid content of rapeseed was simultaneously controlled by the genetic main effects and genotype environment (GE) interaction effects [9]. There was little information about the genetic main effects and their interaction effects from diploid embryo nuclear genes, cytoplasmic /$ see front matter 2004 Elsevier Ireland Ltd. All rights reserved. doi: /j.plantsci

2 44 H.Z. Zhang et al. / Plant Science 167 (2004) genes and diploid maternal plant nuclear genes for OAC of rapeseed. In the present study, analysis was conducted to determine the genetic main effects and their GE interaction effects for OAC of rapeseed. The intents were to clarify the genetic mechanisms including the embryo effects, cytoplasmic effects and maternal effects, as well as their GE interaction effects, and to estimate the heritabilities for OAC. The breeding values of parents were also predicted for OAC in B. napus L. 2. Material and methods 2.1. Field experiments The experiments were carried out by using the diallel design under two environments from 1997 to In the diallel design, eight varieties of B. napus including Youcai 601 (P1), Gaoyou 605 (P2), Huashuang 3 (P3), Yunyou 8 (P4), Zhongyou 821 (P5), Eyouchangjia (P6), Zhong R-888 (P7) and Tower (P8) with different cytoplasms and maternal plant were used as females and males (8 8), which the OAC was ranged from % of total fatty acid content. Total crosses of 28 F 1 sorf 2 s were gained in the experiments. Seeds of the eight parents were sown in October 1997, and the seeds of the parents were self-pollinated and F 1 s were obtained by crossing among eight parents with hand emasculation in the spring of The seeds of parents and F 1 s were both sown on October 7 in 1998 and 1999, and 31-day-old seedlings were individually transplanted at a spacing of 35 cm 30 cm in the experimental farm at Zhejiang University. The experiment was laid out in a randomized block design with three field replications and 24 plants were planted in each plot for parents and F 1. The self-pollinated seeds of parents and F 2 sonf 1 plants from eight plants in the middle part of each plot were derived at maturity, which several anthotaxys were randomly set in the photic bag before anthesis, and the samples of F 1 seeds used for analyzing were obtained during the growing season by using the method of isolated pollination in both environments Data collection OAC of rapeseed was analyzed by gas chromatogram (Japan, Model GC-9A) according to the method proposed by Zhou and Liu [5] and Bouchereau et al. [10]. The oven-dried rapeseeds (1 g) were crushed and transferred to centrifuge tubes in two replications. Thereafter, 1 ml of methylating reagent (methanol/acetyl chloride/benzene, 20:1:4 by volume) was added. The tubes were heated at 60 C for 1.5 h in a water bath. Subsequently, 2 ml of hexane was added and the tubes were centrifuged at 5000 rpm for 5 min. The upper layer was transferred into another tube and the solvent was allowed to evaporate. The methyl esters were dissolved in hexane (250 l) and a 1 l sample was injected into the gas chromatogram and flame ionization detector with operating conditions as follows: temperature (column/injector/detector), 210/240/240 C and carrier gas flow, 30 ml/min. Two parallel times was measured for each sample of OAC, and the errors of the two data were controlled in 0.5%. The statistics of total fatty acid include six kinds fatty acids (palmitic acid, linoleic acid, linoleinc acid, eicosenoic acid and erucic acid) in the measuring. The sum of the six kinds of fatty acid was looked as 100% and OAC (%) was expressed as the percentage of total fatty acid content in rapeseed Statistical analysis The genetic model used was an extension of diploid plant seeds, which could analyze the genetic main effects and GE interaction effects including embryo effects from diploid embryo nuclear genes, cytoplasmic effect from cytoplasmic genes and maternal effects from diploid maternal plant nuclear genes for quantitative traits of rapeseed by using the diallel design with several parents, and MINQUE (0/1) method was also used to estimate variances components [11,12]. Phenotypic variance (V P ) of rapeseed quality traits is composed as follows: V P = V G + V GE + V e = (V A + V D + V C + V Am + V Dm ) + (V AE + V DE + V CE + V AmE + V DmE ) + V e, where V G was genetic main variance, V GE the GE interaction variance, V e the residual variance, V A and V D were embryo additive and dominance variance, V C cytoplasmic variance, V Am and V Dm were maternal additive and dominance variance, V AE and V DE were embryo additive and dominance interaction variance, V CE cytoplasmic interaction variance, V AmE and V DmE were maternal additive and dominance interaction variance, respectively. If OAC of rapeseed was simultaneously controlled by genetic main effects and GE interaction effects of genes from different genetic systems, the total narrow-sense heritability (h 2 ) for OAC could be further partitioned into general heritability (h 2 G ) and interaction heritability (h2 GE ). The conjunction of the components was as follows: h 2 = h 2 G + h2 GE = (h 2 Go + h2 Gc + h2 Gm ) + (h2 GoE + h2 GcE + h2 GmE ), where h 2 Go was embryo general heritability, h2 Gc cytoplasmic heritability, h 2 Gm maternal general heritability, h2 GoE embryo interaction heritability, h 2 GcE cytoplasmic interaction heritability and h 2 GmE maternal interaction heritability. The AUP method was used to predict the genetic main effects including embryo additive main effect (A), cytoplasmic main effect (C), maternal additive main effect (Am) as well as their GE interaction effects (AE, CE and AmE) [13,14].

3 H.Z. Zhang et al. / Plant Science 167 (2004) The standard errors of estimated variances, heritabilities and predicted genetic effects were derived by using the Jackknife method from sampling generation means of entries [14,15]. 3. Results 3.1. Phenotypic means and ranges of the generations for OAC of rapeseed The means and ranges of OAC in rapeseed had large variation among the materials studied (Fig. 1). The range of OAC for eight parents was from to 61.96% in 1999 and to 62.32% in 2000, which was suitable for genetic analysis. The visible differences were also appeared in F 1 and F 2 generations. Since the variation of OAC among different generations was found in different years, the per- formance of OAC in rapeseed could be influenced by environmental conditions (e.g. weather, soil, cultural and management of field) besides of the genetic main effects as well as their GE interaction effects Estimation of variances for OAC of rapeseed The variances of OAC including genetic main variances, GE interaction variances and residual variance were showed in Fig. 2. The results showed that OAC of rapeseed was synchronously controlled by genetic main effects and GE interaction effects, which genetic main effects were more important for this trait since the genetic main variances (V G = V A + V D + V C + V Am + V Dm ) accounted for about 55.2% of the total variances (V G + V GE, V GE = V AE + V DE + V CE + V AmE + V DmE ). All genetic main variance components including V A, V D, V C, V Am and V Dm were significant and the genetic main effects from different genetic systems Fig. 1. Phenotypic distribution of generations for OAC (%). 1999P and 2000P, 1999F 1 and 2000F 1, 1999F 2 and 2000F 2 represented parents, F 1 and F 2 in 1999 and 2000, respectively. Fig. 2. Estimation of genetic main variances and GE interaction variances for OAC of rapeseed. V A : embryo additive main variance, V D : dominance main variance, V C : cytoplasmic main variance, V Am : maternal additive variance, V Dm : maternal dominance variance, V AE : embryo additive interaction variance, V DE : dominance interaction variance, V CE : cytoplasmic interaction variance, V AmE : maternal additive interaction variance, V DmE : maternal dominance interaction variance, and V e : residual variance. All estimates except for V AE were significant at 0.01 level.

4 46 H.Z. Zhang et al. / Plant Science 167 (2004) could affect the performance of OAC. The results of embryo main variances (V A + V D ) accounted for about 55.1% of V G indicated that embryo main effects, especially for embryo additive main effect, were more important in genetic main effects for OAC, while maternal main effects ((V Am + V Dm )/V G = 24.5%) were near to cytoplasmic main effect (V C /V G = 20.3%). For GE interaction variances, the embryo interaction variances (V AE + V DE ) only accounted for 3.8%, but the maternal interaction variances (V AmE + V DmE ) and cytoplasmic interaction variance (V CE ) were larger and accounted for about 56.2 and 39.9% of total V GE, respectively. Therefore, the expression of genes from diploid maternal plant and cytoplasm was easier influenced by the environments than that from diploid embryo. It was suggested by the larger additive and cytoplasmic variances ((V A + V C + V Am )/V G = 83.3%) and their interaction variance (V AE + V CE + V AmE )/V GE = 73.7%) that additive and cytoplasmic effects were more important than other genetic effects for OAC of rapeseed. Better effect for improving OAC of rapeseed could be achieved by selection in early generations. Since residual variance (V e ) was significant, the performance of OAC could be influenced by sampling errors. OAC was mainly controlled by genetic main and GE interaction effects because of the small value of residual variances (V e /(V G + V GE + V e ) = 1.9%) Estimation of heritabilities of rapeseed The estimate of total narrow-sense heritability (h 2 )was 77.5%, with the general heritability (h 2 G ) and interaction heritability (h 2 GE ) accounted for 45.1 and 32.4%, respectively (Fig. 3). For general heritability, h 2 Go from diploid embryo was much lager than h 2 Gc or h2 Gm from cytoplasm or diploid maternal plant, which indicated that embryo general heritability for OAC were more important than the other heritability components. For interaction heritability, h 2 GoE was not found in this study because there was no embryo additive interaction variance (Fig. 2), but h 2 GcE and h2 GmE were both significant for OAC of rapeseed. Since h 2 G and h2 GE were simultaneously found, the select efficiency for OAC could also be influenced by different environments, but the higher value of h 2 for OAC suggested that the selection pursuit could be expected in early generations Prediction of embryo, cytoplasmic and maternal effects of parents In order to improve OAC by using the better parent(s), predicting the genetic merit of parents was also important during the progress of rapeseed quality breeding. Breeding value of each parent for OAC could be predicted from the genetic main effects and GE interaction effects. It was indicated that genetic main effects including A, C and Am and their GE interaction effects including CE and AmE of parents could affect the OAC of offspring. Among eight parents, although the negative maternal additive main effect (Am) and the negative maternal additive interaction effect in 1999 (AmEI) of P7 could decrease OAC, the total breeding values (G t I = A + AEI + C + CEI + Am + AmEI in 1999 and G t II = A + AEII + C + CEII + Am + AmEII in 2000) were still reached 18.6 and 21.0% for P7 (Zhong R-888) in 2 years, respectively (Fig. 4, Table 1). The G t I and G t II for P3 (Huashuang 3) were larger and the total breeding values were 14.5 and 17.4% in 1999 and 2000, respectively. Therefore, P3 (Huashuang 3) and P7 (Zhong R-888) were better than other parents for increasing OAC of offspring in different environments. Since the predicted breeding values for P6 (Eyouchangjia) and P8 (Tower) were both contrary directions in different years, the total genetic effects from these two parents were not stable in different environments. The other parents could reduce the OAC of rapeseed in progenies due to the negative predict value in both years, so these Fig. 3. Estimation of heritability components for OAC of rapeseed. h 2 Go : embryo general heritability, h2 Gc : cytoplasmic heritability, h2 Gm : maternal general heritability, h 2 GoE : embryo interaction heritability, h2 GcE : cytoplasmic interaction heritability and h2 GmE : maternal interaction heritability. All estimates except for h 2 GoE were significant at 0.01 level.

5 H.Z. Zhang et al. / Plant Science 167 (2004) Fig. 4. Predicated total genetic effects (%) for OAC of rapeseed. G t I and G t II referred to the total genetic effect in 1999 and in 2000, respectively. P1: Youcai 601, P2: Gaoyou 605, P3: Huashuang 3, P4: Yunyou 8, P5: Zhongyou 821, P6: Eyouchangjia, P7: Zhong R-888 and P8: tower. Table 1 Predicated genetic effects (%) for OAC of rapeseed Parent Embryo additive effect Cytoplasmic effect Maternal additive effect A AEI AEII C CEI CEII Am AmEI AmEII P P P P P P P P A: embryo additive main effect, AE: embryo additive interaction effect, C: cytoplasm main effect, CE: cytoplasmic interaction effect, Am: maternal additive main effect, AmE: maternal additive interaction effect. Symbol indicates not-availability of predicted effects because of the corresponding zero estimate of variance component in Fig. 2. Significant at 0.05 level. Significant at 0.01 level. Significant at parents were not suitable in high OAC breeding of rapeseed (Fig. 4). 4. Discussion It was well known that the development of rapeseed oil with high OAC was important for both food- and non-food-purposes. The utilization of classical plant breeding techniques was still a basal method for improving fatty acid composition of rapeseed. It is useful for breeders to improve the efficiency in breeding following the genetic mechanism(s) of quality traits of rapeseed. For diploid crops, such as B. napus, the main part of seed is embryo (cotyledon) and the seed development is based on the photosynthetic product from maternal plant. Chloroplast and/or mitochondrion DNA existing in cytoplasm could also affect the quality of rapeseed. Therefore, quality traits of rapeseed might be simultaneously controlled by the genes from different genetic systems including diploid embryo nuclear genes, diploid maternal plant nuclear genes and cytoplasmic genes. The results of this experiment showed that OAC of rapeseed could be affected by different genetic system effects, and the embryo genetic effects were found to be the main cause for the variation of this trait. The cytoplasmic and maternal effects were also important for the performance of OAC and could not be ignored in the rapeseed quality breeding. Rapeseed growing in field can be influenced by genetic effects and environmental conditions. The climatic factors and cultural differences were the main causes which could affect the expression of genes for the quantitative traits of rapeseed under different environments. The experiments are, therefore, better to do in different environments because of

6 48 H.Z. Zhang et al. / Plant Science 167 (2004) the diversity especially for the climatic conditions. In this experiment, the temperature at flowering and maturing periods of rapeseed in 2000 was much higher than that in 1999, but the rainfall was more in Otherwise, some water and manure management differences of field might be existed between 2 years. Hence, the GE interaction effects can cause genetic differences of seed quality traits among environments. Understanding the magnitude of genetics main effects and GE interaction effects is very useful for improving the efficiency of quality breeding and is also helpful for breeders to select the better lines (varieties) of rapeseed which can be more steady in different environments. The results in this research indicated that the genetic main effects and GE interaction effects from different genetic systems were all important for OAC. It should be pay more attention to the GE interaction effects during the high OAC breeding of rapeseed. In quality breeding, it is useful for rapeseed breeders to improve OAC through selecting better parent(s) by knowing the genetic merit of parents besides understanding inheritance of quality trait. In this experiment, some parents such as Huashuang 3 had better breeding value for improving OAC of offspring due to the positive predict value and the stable genetic effects in different environments (years), so it could be used as parent in quality breeding of rapeseed. Acknowledgements The project was financially supported from Foundation for University Key Teacher by the Ministry of Education of China and by the 151 Program for the Talents of Zhejiang Province. The authors are grateful to Prof. Zhu for providing the analyzing software. References [1] C. Mollers, Development of high oleic acid oilseed rape, in: Presentation at the 8th International Conference for Renewable Resources and Plant Biotechnology, NAROSSA, Magdeburg, 2002, pp [2] M.E. Cartea, M. Migdal, A.M. Galle, G. Pelletier, P. Guerche, Comparison of sense and antisense methodologies for modifying the fatty acid composition of Arabidopsis thaliana oilseed, Plant Sci. 136 (1998) [3] Z.P. Kondra, S.T. Tsunoda, Inheritance of oleic, linoleic and linolenic acids in seed oil of rapeseed (Brassica napus L.), Can. J. Plant Sci. 55 (1975) [4] R. Ecker, Z. Yaniv, Gentic control of fatty acid composition in seed oil of Sinapis alba L., Euphyica 69 (1993) [5] Y.M. Zhou, H.L. Liu, Inheritance of fatty acid in rapeseed (B. napus), Acta Agronomica Sinica 13 (1987) [6] J.L. Chen, W.D. Beversdorf, Fatty acid inheritance in microsporederived population of spring rapeseed (Brassica napus L.), Ther. Appl. Genet. 80 (1990) [7] S. Pleines, W. Friedt, Breeding for improved C18-fatty acid composition in rapeseed (Brassica napus L.), Fat Sci. Technol. 90 (1988) [8] A. Schierholt, H.C. Becker, Environmental variability and heritability of high oleic acid content in winter oilseed rape (Brassica napus L.), Plant Breed. 120 (2001) [9] C.H. Shi, H.Z. Zhang, J.G. Wu, C.T. Li, Y.L. Ren, Genetic and genotype (environment interaction effects analysis for erucic acid content in rapeseed (Brassica napus L.), Euphytica 130 (2003) [10] A. Bouchereau, N. Clossais-Besnard, A. Bensaoud, L. Leport, M. Renard, Water stress effects on rapeseed quality, Eur. J. Agron. 5 (1996) [11] J. Zhu, B.W. Weir, Analysis of cytoplasmic and maternal effects. I. A genetic model for diploid plant seeds and animals, Theor. Appl. Genet. 89 (1994) [12] J. Zhu, Analytic methods for seed models with genotype environment interactions, Acta Genetica Sinca 23 (1996) [13] J. Zhu, Methods of predicting genotype value and heterosis for offspring of hybrids, J. Biomath. 8 (1) (1993) [14] J. Zhu, B.W. Weir, Diallel analysis for sex-linked and maternal effects, Theor. Appl. Genet. 92 (1996) 1 9. [15] R.G. Miller, The Jackknife, a review, Biometrika 61 (1974) 1 15.