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Identification of Quantitative Trait Loci for Fatty Acid Content in Brown Rice (Oryza sativa L.)
Plant Breeding and Biotechnology 2018;6:444-453
Published online December 31, 2018
© 2018 Korean Society of Breeding Science.

Su Jang1, and Joong Hyoun Chin2,*

1Department of Plant Science, Plant Genomics and Breeding Institute, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul 08826, Korea, 2Department of Integrative Bio-industrial Engineering, Sejong University, Seoul 05006, Korea
Corresponding author: *Joong Hyoun Chin, jhchin@sejong.ac.kr, Tel: +82-2-6935-3897
Received November 16, 2018; Revised November 18, 2018; Accepted November 18, 2018.
This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract

The rice bran oil contained in brown rice is composed of highly valued ingredient. Improving the content of unsaturated fatty acids in rice seed, such as oleic acid, linoleic acid, and α-linolenic acid, would provide more benefit to human health. Fatty acid content is quantitative trait controlled by multiple genes. We have utilized high-density SNP data from highly advanced breeding populations to identify QTLs for fatty acid contents in brown rice. Here, we identified 51 major QTLs (M-QTLs) and 25 epistatic QTLs (EpQTLs) related to eleven fatty acid contents. Eight and four M-QTLs were pleiotropically associated with the content of different fatty acids in MT-RILs and DT-RILs, respectively. Total effect of M-QTLs for palmitic acid (16:0), oleic acid (18:1), and linoleic acid (18:2), could explain phenotypic variations of 36.7%, 63.7%, and 41% in MT-RILs, respectively. Alpha-linolenic acid which is important for a human’s health could be explained phenotypic variation of 15.7% by six M-QTLs. These QTLs identified in this study can be used to improve nutritious content in rice breeding programs.

Keywords : Rice, Fatty acid, QTL, Brown rice, Unsaturated fatty acid
INTRODUCTION

Brown rice itself is consumed as a type of healthy food in many countries. The oil contained in rice bran is composed of many useful healthy chemical compounds. After milling, the rice bran, as a form of by-products, is used to extract high-quality oil. All plant seeds contain seed storage lipids in the form of triacylglycerol (TAG). TAG contains a backbone of the esterified fatty acids. Three fatty acids including Palmitic acid (16:0; Pal), Oleic acid (18:1 Δ 9, n-9;Ole), and Linoleic acid (18:2 Δ9,12, n-6; Lin) are major fatty acid component of rice seed (Yasumatsu and Moritaka 1964; Resurreccion and Juliano 1975).

The optimum level of linoleic/α-linolenic acid ratio (Lin/Ali ratio, LA) for the growth of the human diet, especially for infants and children, was estimated in various ways (Jensen et al. 1997; Hejr et al. 2017). On the other hand, due to the benefit of high level of oleic acid (Ole) content to cardiovascular disease (Lopez-Huertas 2010) and etc., a trial was set to increase Ole by editing the OsFAD2-1 gene, which converts Ole to Lin (Abe et al. 2018).

By the help of genomics advancements, QTLs associated with the lipid contents of some plants were reported in many plants, including rice (Panthee et al. 2006; Barker et al. 2007; Haddadi et al. 2010; Sarvamangala et al. 2011; Ying et al. 2012). The metabolic pathway of plant lipids in seeds has been studied, and the co-locations of the identified QTLs/genes have been reported in Arabidopsis, maize, sunflower, rape, and rice (Pérez-Vich et al. 2002; Hobbs et al. 2004; Yang et al. 2010; Smooker et al. 2011; Ying et al. 2012). However, studies related with lipid/fatty acid QTLs, utilizing sequence-based genotyping system from highly advanced breeding populations such recombinant inbred lines in rice, have rarely been conducted.

In this study, two indica-japonica recombinant inbred lines (RILs) were employed to identify the additive and epistatic QTLs associated with fatty acid contents in rice. With the results from this study, the breeding for brown rice containing high levels of healthy fatty acid content could be facilitated by molecular breeding approaches.

MATERIALS AND METHODS

Plant materials and growing in the field

Two recombinant inbred lines (RILs) derived from intersubspecific (indica-japonica) crosses by the single-seed descendant method were employed in this study (Yoo 2017). A total of 134 RILs derived from a cross between Dasanbyeo (Tongil type, indica) and TR22183 (temperate japonica) (DT-RILs), and 155 RILs developed from Milyang 23 (Tongil type, indica) and Tong 88-7 (temperate japonica) (MT-RILs) by Seoul National University in Korea were used in this study. The plant materials, together with the parental lines, were grown at the Seoul National University Experimental Farm in Suwon in 2015. The plants were transplanted to one seedling per hill at a planting density of 30 × 15 cm. The two RILs and their parents were cultivated under regularly irrigated and fertilized conditions (N–P2O5–K2O = 100–80–80 kg/ha).

Fatty acid preparation

The extraction of lipids in whole grain brown rice and milled rice followed the method of Folch et al. (1957). The extracted lipid was methyl esterized to the method of Lepage and Roy (1986) and analyzed. The Fatty acid analysis was conducted at NICEM of Seoul National University. The analyzed fatty acids are as follows. Unsaturated fatty acids: C18:1 (Oleic acid: Ole); C18:2 (Linoleic acid: Lin); C18:3 (α-Linolenic acid: Ali); and C20:1 (cis-11-Eicosenoic acid: Eic). Saturated fatty acids: C14:0 (Myristic acid: Myr); C16:0 (Stearic acid: Ste); C18:0 (Palmitic acid: Pal); C20:0 (Arachidic acid: Ara); C22:0 (Behenic acid: Beh); C23:0 (Tricosanoic acid: Tri); and C24:0 (Lignoceric acid: Lig).

Linkage map construction and QTL analysis

Genomic DNA was extracted from 2-cm fresh leaves from seedlings at the three-leaf stage, using cetyltrimethylammonium-bromide (CTAB) (Murray and Thompson 1980). For DT-RILs, 384-plex SNP genotyping, using the Illumina GoldenGate Assay, was performed using VeraCode technology on the BeadXpress reader (Fan et al. 2003; Thomson et al. 2012). The DNA of MT-RILs were genotyped by genotyping-by-sequencing (GBS). A GBS library was prepared with the restricted enzyme ApeKI, as described by Elshire et al. (2011). For the accurate variation calling from our RIL population of intersubspecific cross, we prepared subset of rice reference genome with ApeKI restriction sites that are target genomic regions for GBS genotyping with IRGSP 1.0 pseudomolecule. We in silico-searched the ApeKI sites in the reference genome sequences and split it into fragments. The subset of genome fragments smaller than 2kb were collected. The subset of genome was indexed and the GBS short reads were mapped using software BWA (Li and Durbin 2009). The genotypes of RIL population were determined into homo- and heterozygous genotypes by software SAMtools with default parameter (Li et al. 2009). For cross-validation, the parents, genomic DNA of M23 and T887, were fully sequenced using the HiSeq2000 platform (Illumina, Inc., San Diego, CA) to compare allele contents for each locus with GBS results. Only the called SNPs of GBS results which matched with resequencing results of the parents, M23 and T887, were chosen for the following analyses. Actually, The same linkage map of DT-RILs and MT-RILs were employed to conduct the QTL analysis (Yoo 2017). QTL analysis was carried out using the software ICIMapping 4.1 (Meng et al. 2015).

RESULTS

Milyang 23 (M23) contained more fatty acids than Tong 88-7 (T887) in three unsaturated fatty acids (Oleic acid, Linoleic acid, and α-Linolenic acid) and in four saturated fatty acids (Palmitic acid, Stearic acid, Arachidic acid, and Behenic acid) (Table 1). The ratio of unsaturated fatty acids to saturated fatty acids (US) of M23 was higher than that of T887. However, the ratio of linoleic acid to α-linolenic acid (LA) of two parents were not different. Dasanbyeo (DS) contained more unsaturated and saturated fatty acids than those of TR22183 (TR). Interestingly, the Tongil-type or temperate indica parents contained more fatty acids than those of japonica parents in brown rice. However, the amount of Ole, Ste, and Beh of TR was not different from that of DS. The distribution of fatty acids from the two RILs showed a normal distribution, except for Ali and Tri of DT-RILs.

All fatty acids showed positive correlations with the different significance level (Fig. 1). The correlation between the fatty acids in DT-RILs was more obvious than those of MT-RILs. Ole, Eic and US showed significant correlations in both the RILs.

QTLs for fatty acid contents

In MT-RILs, there were a total of 22 major QTLs (M-QTLs) for unsaturated fatty acids and 24 for saturated fatty acids (Table 2). The QTLs for total unsaturated fatty acids (Unsat) were identified on chromosomes 1, 5, 7, and 9, which are co-located with QTLs for Ole (qOle1.1, qOle5.1, qOle7.1, and qOle9.1), Eic (qEic5.1 and qEic9.1), and Lin (qLin7.1). The QTL for Lin on chromosome 6 (qLin06.1) showed the largest phenotype variation explained (PVE) (26.22%) among all the unsaturated fatty acid QTLs. The QTL for total saturated fatty acids (Sat) were identified on chromosome 7 (qSat07.1), which is co-located with QTLs for Myr (qMyr07.2) and Pal (qPal07.1). Interestingly, qSat07.1 and qUnsat07.1 seemed to be very closely located (Fig. 2A). A total of 10 fatty acid QTLs were identified on chromosome 7.

In DT-RILs, only one QTL for unsaturated fatty acid and four QTLs for saturated fatty acids were identified for Ali (qAli03.1). The qSat01.1 for total saturated fatty acids was co-located with qAra01.1 for Ara. All the QTLs for saturated fatty acids were closely located on chromosome 1 (Fig. 2B).

In MT-RILs, only one QTL for Ole was identified on chromosome 2 (EpOle0202.1), which was co-located with EpUnsat02.1 (Fig. 2A). Two interacting loci of EpOle02.1 were closely linked with the two QTLs for Eic (qEic02.1 and qEic02.2). The other two epistatic QTL (EpQTLs) for Sat seemed not to be linked with any other major QTLs. On the other hand, a total of 23 EpQTLs for Ali and one EpQTL for Ste were identified in DT-RILs (Fig. 2B). Not like EpQTLs in MT-RILs, many small EpQTLs with small PVE in DT-RILs, however the total PVE of all the EpQTLs were up to 44.9% for Ali (Table 3).

DISCUSSION

The rice bran oil contained in brown rice is beneficial to human health. To improve palatability, most rice consumption is from milled rice, which results in rice bran, including seed coat and rice embryo to be discarded. However, with the nutritious effect of rice bran being intensively studied, whole grain brown rice consumption is increasing, especially in Northeast Asian countries. Rice bran crude extract became famous for being a highly valued ingredient in high price cosmetics and supplementary nutrition products.

Three fatty acids are majorly found In rice seed: two unsaturated acids (Oleic acid and linoleic acid) and one saturated acid (palmitic acid) (Yasumatsu and Moritaka 1964; Resurreccion and Juliano 1975). In our study, the Ole of the parents of MT-RIL showed difference, however that of DT-RIL did not (Table 1). The total PVE of M-QTLs and EpQTLs for Ole in MT-RILs was 75.41%. On the contrary, there was no QTL for Ole identified in DT-RILs (Table 4). In the same way, the total PVE of Lin of MT-RIL was 40.96%, but no QTL was identified in DT-RIL. The PVE of the most common saturated fatty acid, Pal, was also only estimated in MT-RIL, as 36.7%. So, the identification of QTLs for major fatty acids were possible only in MT-RIL, in this study. It was mainly due to the level of difference of each QTL between parents and the presence (Table 1).

The correlation between fatty acids in MT- and DT-RILs showed similar ways, and it was found for the DT-RILs to have higher degrees of positive correlations (Fig. 1). It implies that the genetic structure regarding fatty acids were sharing genes associated with fatty acid metabolism.

In MT-RILs, fatty acid QTLs were identified through all chromosomes except for chromosome 8, while those of DT-RILs were only on chromosomes 1 and 3 (Fig. 2). However, the DT-RIL showed many EpQTLs for Ali on all chromosomes. Although the total PVE of all EpQTLs for Ali of DT-RIL is 45.88%, it is cautious to conclude that DT-RIL has that much of a complicated genetic structure for Ali. So, the isolation of QTLs for Lin and Ali would be more feasible for further molecular breeding approaches.

Ying et al. (2012) reported 29 QTLs for fatty acids and oil content. Comparing with our study, it seems that only the QTL for Ole on chromosome 1 is identified in our study (qOle01.2), too. All other QTLs were identified for the first time in our study. It is also difficult to compare directly, due to the limit of molecular markers and the fact that F2:3 early generation population was used in their study.

TAGs are major form of storage lipids in plant seed. Plant TAG synthesis can be separated in three phases, including (1) fatty acid synthesis, (2) modification of fatty acids, and (3) incorporation of fatty acids into TAGs (Leskinen 2010). Several candidate genes involved in lipid metabolism were located within nine QTL regions controlling Ole, Lin, and Ali in MT-RILs. Especially, genes encoding phospholipase were found in qOle01.1 and qOle06.1, which had pleotropic effect for different fatty acids. Phospholipases are enzymes which catalyze hydrolysis of phospholipids, including phosphatidylcholine (PC). PC is the main substrate for the desaturation of oleic acid to linoleic acid and α-linolenic acid. In addition, fatty acid from PC can be used to TAG synthesis (Leskinen 2010). Since the function of candidate genes in qOle01.1 and qOle06.1 affects the contents of Ole and Lin, these genes could be regarded as strong candidates for target traits.

In both RILs, the total unsaturated fatty acid contents were far lower than those of the parents (Table 1). It is interesting to know that no RIL contained more amount of total Unsat than those of the parents. However, the range of each fatty acid showed that a few lines contained high levels of unsaturated fatty acids. Thus, the pyramiding of favorable QTLs for each unsaturated fatty acid should be combined by molecular marker approach to improve the total content of unsaturated fatty acid of rice grain.

ACKNOWLEDGEMENTS

This study was supported by a grant from the Next-Generation BioGreen 21 Program (no. PJ01319603) of the Rural Development Administration, Korea. This research was also supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (no. 2017R1D1A1B04034862).

Figures
Fig. 1. Correlation matrix plot between fatty acid contents in MT-RILs and DT-RILs. Correlation matrix plot represents Pearson’s correlation coefficient in 155 MT-RILs (A) and 134 DT-RILs (B). Clustering is conducted by FPC function of corrplot package in R. Color scale shows the range from lowest correlation (dark red) to highest correlation (dark green) coefficient.
Fig. 2. The results of QTL analysis for fatty acid contents in MT-RILs and DT-RILs. Circos diagrams show QTL and Epistatic network in MT-RILs (A) and DT-RILs (B). The vertical line denotes M-QTLs (LOD > 2.5) position in chromosome. Inner connection represents an interaction between EpQTLs (LOD > 5).
Tables

Summary statistics of fatty acid contents of the two RILs and their parents.

Unsaturated fatty acid Saturated fatty acid USx)



C18:1 Oley) C18:2 Lin C18:3 Ali C20:1 Eic Unsat LA C14:0 Myr C16:0 Pal C18:0 Ste C20:0 Ara C22:0 Beh C23:0 Tri C24:0 Lig Sat US (unsat/sat)
Milyang 23 10.63**w) 9.98** 0.35* 0.12ns 21.08** 28.63ns 0.13** 6.27** 0.62** 0.18* 0.11* 0.02ns 0.25ns 7.61** 2.77**
Tong 88-7 9.50 8.66 0.30 0.14 18.61 28.41 0.22 5.84 0.50 0.15 0.08 0.02 0.24 7.09 2.62
MT-RILs (n = 155)
 Range 6.727–14.93 7.683–15.23 0.262–0.567 0.078–0.205 6.563–10.67 19.66–39.31 0.066–0.39 5.266–8.579 0.473–1.454 0.122–0.345 0.064–0.186 0.012–0.025 0.189–0.445 16.18–29.07 2.137–3.31
 Mean 10.85 10.79 0.39 0.14 8.47 28.38 0.20 6.76 0.78 0.21 0.13 0.02 0.33 22.17 2.62
 SDz) 1.77 1.29 0.05 0.03 0.82 4.07 0.06 0.65 0.18 0.05 0.02 0.00 0.05 2.58 0.21
 Skewness 0.01 0.62 0.38 0.16 0.28 0.50 0.48 0.35 0.76 0.69 0.19 0.19 −0.18 0.17 0.25
 Kurtosis −0.63 0.78 0.39 −0.70 −0.41 0.06 0.22 −0.19 0.59 0.69 0.00 −0.14 −0.37 −0.27 0.27
Dasanbyeo 11.25ns 9.88** 0.38* 0.12ns 21.63* 25.85ns 0.20* 6.64** 0.69ns 0.21* 0.11ns 0.02ns 0.25ns 8.12** 2.66**
TR22183 11.80 8.35 0.33 0.14 20.62 25.28 0.24 5.93 0.67 0.17 0.11 0.02 0.26 7.42 2.78
DT-RILs (n = 134)
 Range 7.443–19.13 7.307–13.28 0.243–0.872 0.088–0.239 6.12–11.93 10.26–34.79 0.108–0.441 4.813–9.44 0.403–1.326 0.128–0.346 0.083–0.183 0.011–0.042 0.165–0.459 15.1–32.45 2.137–3.127
 Mean 11.87 9.91 0.38 0.14 8.39 26.37 0.25 6.67 0.77 0.21 0.12 0.02 0.31 22.31 2.66
 SD 2.01 1.13 0.07 0.03 0.96 3.55 0.06 0.75 0.16 0.05 0.02 0.00 0.05 2.85 0.19
 Skewness 0.96 0.37 2.29 0.66 0.79 −0.70 0.69 0.77 0.85 0.71 0.60 1.65 0.15 0.88 −0.48
 Kurtosis 1.62 −0.01 13.40 0.36 1.14 2.25 0.66 1.25 1.55 0.21 0.06 8.47 0.18 1.80 0.13

SD: standard deviation,

Ole: oleic acid, Lin: linoleic acid, Ali: α-linolenic acid, Eic: cis-11-Eicosenoic acid, Unsat: total content of all unsaturated fatty acid, LA: the ratio of linoleic acid to α-linolenic acid, Myr: myristic acid, Pal: palmitic acid, Ste: Stearic acid, Ara: arachidic acid, Beh: behenic acid, Tri: tricosanoic acid, Lig: lignoceric acid, Sat: total content of all saturated fatty acid,

US, The ratio of total content of all unsaturated fatty acids to saturated fatty acids,

ns, *, ** and *** stand for not significant at the 0.05 probability level, significant at the 0.05, 0.01, and 0.001 probability level, respectively.


Major QTLs for fatty acid contents in two recombinant inbred lines.

RILs Fatty acids QTLsz) Chry) Left marker Right marker LODx) PVE (%)w) Av)
MT-RILs Unsaturated fatty acid qAli07.1 7 07_16952465 07_17096594 5.68 15.67 −0.02
qEic02.1 2 02_26456447 02_26465513 4.27 5.62 0.01
qEic02.2 2 02_6091905 02_6121376 8.07 11.18 −0.01
qEic03.1 3 03_31064070 03_31115927 6.37 8.48 0.01
qEic05.1 5 05_23196471 05_23432616 9.97 14.48 −0.01
qEic09.1 9 09_22183881 09_22226710 4.70 6.16 −0.01
qEic10.1 10 10_18055887 10_19521655 6.08 10.04 0.01
qLin06.1 6 06_24427292 06_24449197 12.20 26.22 0.66
qLin07.1 7 07_21597703 07_22087171 7.42 14.74 −0.47
qOle01.1 1 01_29280527 01_29469791 5.02 6.10 0.41
qOle01.2 1 01_33869585 01_33924337 5.20 6.22 0.41
qOle05.1 5 05_23196471 05_23432616 12.07 16.59 −0.68
qOle06.1 6 06_23963143 06_24107686 7.87 9.84 −0.53
qOle07.1 7 07_21299763 07_21346892 9.45 12.06 −0.58
qOle09.1 9 09_22183881 09_22226710 6.02 7.31 −0.46
qOle10.1 10 10_18055887 10_19521655 4.44 5.54 0.39
qLA01.1 1 01_5594605 01_5760962 7.45 8.15 1.32
qLA05.1 5 05_16329242 05_16344049 5.50 5.68 −1.12
qLA06.1 6 06_24427292 06_24449197 7.89 8.71 1.42
qLA06.2 6 06_4884640 06_5113827 8.29 9.00 −1.40
qLA07.1 7 07_11635687 07_14689865 15.73 19.36 2.02
qLA07.2 7 07_6865498 07_6947386 7.83 8.40 −1.34
qUnsat01.1 1 01_30129457 01_30253847 4.11 7.25 0.66
qUnsat05.1 5 05_23526681 05_23611087 3.62 6.05 −0.61
qUnsat07.1 7 07_21346892 07_21597703 11.07 21.22 −1.14
qUnsat09.1 9 09_22183881 09_22226710 4.82 8.20 −0.73
Saturated fatty acid qAra01.1 1 01_22127065 01_22283397 9.19 14.58 0.02
qAra05.1 5 05_27025300 05_27067823 5.05 7.53 0.01
qAra07.1 7 07_28506119 07_28650972 6.16 9.49 −0.02
qAra07.2 7 07_4536799 07_4739001 3.83 5.60 0.01
qAra09.1 9 09_18928987 09_18944615 4.68 6.95 −0.01
qBeh03.1 3 03_13093920 03_15647398 5.96 15.23 −0.01
qLig07.1 7 07_7673973 07_8082302 4.14 8.83 0.02
qMyr01.1 1 01_15854138 01_20716165 6.63 4.21 −0.01
qMyr03.1 3 03_5274774 03_5408863 6.70 4.28 0.01
qMyr06.1 6 06_4843357 06_4884640 6.15 3.63 −0.01
qMyr07.1 7 07_11635687 07_14689865 24.00 19.11 0.03
qMyr07.2 7 07_21346892 07_21597703 3.92 2.28 −0.01
qMyr11.1 11 11_26453505 11_26528819 39.94 41.90 0.04
qMyr12.1 12 12_1076787 12_1492123 4.10 2.41 0.01
qPal04.1 4 04_2047622 04_2459932 4.97 9.76 0.20
qPal07.1 7 07_23897595 07_24485301 12.03 26.95 −0.33
qSte01.1 1 01_22127065 01_22283397 6.52 7.78 0.05
qSte01.2 1 01_29469791 01_29676099 8.24 10.15 0.06
qSte03.1 3 03_10486128 03_10615953 4.45 5.22 −0.05
qSte03.2 3 03_6994218 03_7090130 5.66 6.74 −0.05
qSte06.1 6 06_25231934 06_25351600 3.72 4.25 −0.04
qSte07.1 7 07_8082302 07_8752549 12.19 16.26 0.08
qSte09.1 9 09_18928987 09_18944615 3.72 4.26 −0.04
qSte11.1 11 11_26453505 11_26528819 4.57 5.34 −0.04
qSat07.1 7 07_23897595 07_24485301 7.00 18.61 −0.33
Ratio of unsaturated/saturated qUS03.1 3 03_7456990 03_7710680 7.24 9.33 0.07
qUS04.1 4 04_2047622 04_2459932 5.68 6.90 −0.05
qUS04.2 4 04_32144327 04_32171508 4.23 5.03 −0.05
qUS05.1 5 05_23196471 05_23432616 10.08 13.28 −0.07
qUS07.1 7 07_5803738 07_5977720 5.56 6.83 −0.05
qUS09.1 9 09_22226710 09_22810010 4.57 5.69 −0.05
qUS11.1 11 11_26453505 11_26528819 10.96 14.55 −0.08
DT-RILs Unsaturated fatty acid qAli03.1 3 C3_15747219 C3_16950533 8.70 9.35 −0.23
Saturated fatty acid qAra01.1 1 C1_19347439 C1_21202056 4.57 14.86 0.02
qBeh01.1 1 C1_17606402 C1_19347439 5.30 16.60 0.01
qSte01.1 1 C1_17606402 C1_19347439 4.46 11.98 0.06
qTri01.1 1 C1_17276596 C1_17606402 46.56 23.72 0.01
qSat01.1 1 C1_19347439 C1_21202056 4.19 11.93 0.34

The abbreviations are same in Table 1,

Chromosome,

Logarithm of odds ratio. LOD threshold more than 2.5 was used to detect QTLs related to fatty acid traits,

Percent of the phenotypic variance explained,

Additive effect. Positive and negative values of additive effect indicate effect contributed by Milyang23 or Dasanbyeo (female parent) and Tong88-7 and TR22183 (male parent) allele, respectively.


Epistatic QTLs for fatty acid contents in two recombinant inbred lines.

RILs EpQTLsw) Loci i Loci j LODz) PVE (%)y) A ijx)


Chrv) Flanking markers Chr Flanking markers
MT-RILs EpOle0202.1 2 02_5116309 – 02_5276878 2 02_26923436 – 02_27114074 7.32 11.75 −0.47
EpSat0406.1 4 04_18049014 – 04_18548596 6 06_19206019 – 06_21373042 5.46 15.71 −0.32
EpSat0808.1 8 08_1995235 – 08_2149557 9 09_15884941 – 09_15961709 5.45 12.86 0.28
EpUnsat0202.1 2 02_4448864 – 02_4555050 2 02_26923436 – 02_27114074 5.18 10.62 −0.73
DT-RILs EpAli0101.1 1 C1_13301985 – C1_15181352 1 C1_19347439 – C1_21202056 7.8149 1.8825 −0.11
EpAli0202.1 2 C2_10272049 – C2_11416702 2 C2_15475666 – C2_17164264 9.291 1.9067 −0.12
EpAli0303.1 3 C3_7472641 – C3_9795740 3 C3_11128609 – C3_13300620 9.1631 1.9581 −0.13
EpAli0103.1 1 C1_2258237 – C1_3711264 3 C3_15747219 – C3_16950533 9.3528 1.8701 0.12
EpAli0203.1 2 C2_18346536 – C2_19700734 3 C3_15747219 – C3_16950533 7.6902 1.9287 −0.11
EpAli0304.1 3 C3_15747219 – C3_16950533 4 C4_23105648 – C4_23695498 9.5615 1.9196 0.12
EpAli0404.1 4 C4_29330690 – C4_30085669 4 C4_30085669 – C4_33349512 7.3736 1.9484 −0.13
EpAli0305.1 3 C3_15747219 – C3_16950533 5 C5_2450054 – C5_3680398 9.2147 1.9127 0.12
EpAli0505.1 5 C5_21558194 – C5_22591851 5 C5_23962325 – C5_28003657 6.8009 2.0356 −0.13
EpAli0306.1 3 C3_15747219 – C3_16950533 6 C6_243274 – C6_7096652 7.8865 2.0178 −0.11
EpAli0606.1 6 C6_7096652 – C6_8725792 6 C6_8725792 – C6_10550728 5.7685 1.8682 −0.12
EpAli0307.1 3 C3_11128609 – C3_13300620 7 C7_2535000 – C7_3318320 9.1484 1.951 −0.13
EpAli0707.1 7 C7_21783591 – C7_23146522 7 C7_23146522 – C7_23513073 7.1038 2.0815 −0.13
EpAli0808.1 8 C8_5846154 – C8_6227953 8 C8_8424668 – C8_9201609 7.8396 1.8937 −0.11
EpAli0308.1 3 C3_11128609 – C3_13300620 8 C8_23650107 – C8_24758913 9.2993 1.9783 0.13
EpAli0909.1 9 C9_14862495 – C9_16324534 9 C9_17348492 – C9_19338543 5.2015 1.9493 −0.11
EpAli0309.1 3 C3_15747219 – C3_16950533 9 C9_20834296 – C9_21464824 7.8601 1.8779 0.12
EpAli0310.1 3 C3_15747219 – C3_16950533 10 C10_19348858 – C10_21045327 8.1436 1.9033 −0.12
EpAli0311.1 3 C3_15747219 – C3_16950533 11 C11_18881908 – C11_20243512 7.8439 1.8871 −0.12
EpAli1111.1 11 C11_21814416 – C11_22706944 11 C11_22706944 – C11_28435768 5.7655 1.9262 −0.12
EpAli1212.1 12 C12_20783152 – C12_21809823 12 C12_21809823 – C12_24602108 7.5372 1.9922 −0.11
EpAli0312.1 3 C3_16950533 – C3_23013821 12 C12_21809823 – C12_24602108 9.9041 1.9748 0.13
EpAli0512.1 5 C5_23962325 – C5_28003657 12 C12_21809823 – C12_24602108 6.3313 2.273 0.13
EpSte0101.1 1 C1_39992906 – C1_40879967 1 C1_41280448 – C1_42541933 6.633 14.0726 −0.13

Logarithm of odds ratio, LOD threshold more than 2.5 was used to detect QTLs related to fatty acid traits,

percent of the phenotypic variance explained,

additive-by-additive effect estimated from the additive effect of each loci (i and j),

epistatic QTLs,

chromosome,

percent of the phenotypic variance explained.


Total phenotype variation explained by additive and epistatic QTLs associated with fatty acid contents.

RILsz) fatty acids Traitx) M-QTLy) EpQTLx) Total



No. PVE (%)w) No. PVE (%) No. PVE (%)
MT-RILs Unsaturated Ali 1 15.67 0 0 1 15.67
Eic 6 55.97 0 0 6 55.97
Lin 2 40.96 0 0 2 40.96
Ole 7 63.66 1 11.75 8 75.41
Lin/Ali 6 59.29 0 0 6 59.29
Unsat 4 42.72 1 10.62 5 53.34
Saturated Ara 5 44.14 0 0 5 44.14
Beh 1 15.23 0 0 1 15.23
Lig 1 8.83 0 0 1 8.83
Myr 7 77.83 0 0 7 77.83
Pal 2 36.7 0 0 2 36.7
Ste 7 52.22 0 0 7 52.22
Sat 1 18.61 2 28.56 1 47.17
DT-RILs Unsat/sat unsaturated saturated US 7 61.62 0 0 7 61.62
Ali 1 9.35 23 44.94 24 54.29
Ara 1 14.86 0 0 1 14.86
Beh 1 16.6 0 0 1 16.6
Ste 1 11.98 1 14.07 2 26.05
Tri 1 23.72 0 0 1 23.72
Sat 1 11.93 0 0 1 11.93

Recombinant inbred lines,

major QTLs,

epistatic QTLs,

percent of the phenotypic variance explained,

see abbreviations in Table 1.


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