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Acta Agronomica Sinica ›› 2023, Vol. 49 ›› Issue (5): 1197-1210.doi: 10.3724/SP.J.1006.2023.24105

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Preliminary exploration of the role of LncRNA in the ecotype differentiation of Brassica napus L.

YANG Tai-Hua(), YANG Fu-Quan, GAO Geng-Dong, YIN Shuai, JIN Qing-Dong, XU Lin-Shan, KUAI Jie, WANG Bo, XU Zheng-Hua, GE Xian-Hong, WANG Jing(), ZHOU Guang-Sheng   

  1. College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, Hubei, China
  • Received:2022-04-27 Accepted:2022-09-05 Online:2023-05-12 Published:2022-09-15
  • Contact: *E-mail: wangjing@mail.hzau.edu.cn
  • Supported by:
    National Key Research and Development Program of China “Physiological Basis and Agronomic Management for High-quality and High-yield of Field Cash Crops”(2018YFD1000900)

Abstract:

Brassica napus L. is one of the important oil crops in China. Based on its different requirements for vernalization temperature during flower transition, Brassica. napus can be divided into three ecotypes (spring, semi-winter, and winter ecotype). Previous studies found that long non-coding RNA (LncRNA) can regulate gene expression level in multiple levels and participate in the regulation of plant growth and development. In Arabidopsis thaliana, LncRNA can regulate flowering time by regulating the expression of genes related to vernalization pathway. In this study, to preliminarily explore the ecotype differentiation and adaptive formation among the three ecotypes of Brassica napus, three different ecotypes of Brassica. napus were used as the materials and high-throughput sequencing technology was used to sequence mRNA and LncRNA in seeding leaves. The GO and KEGG enrichment analysis of co-differential expression genes of three ecotypes of Brassica napus showed that there were a lot of differences in the anabolism of basic compounds, especially lipid compounds. A total of 3775 LncRNA sequences were identified from the three ecotypes of Brassica napus, among which 285 LncRNA were differentially expressed in two or three ecotype combinations, and 1517 candidate target genes were involved. The candidate target genes of these differential expression LncRNA were also enriched in large number of anabolism related pathways of basic compounds. Based on conjoint analysis of mRNA and LncRNA, we predicted a putative regulatory network in the flowering genes, included eight flowering time genes and 23 LncRNA, which were involved in the regulation of temperature and light signal pathways. By analyzing the location of QTLs for important agronomic traits in B. napus, we found that about 90% of LncRNA and QTLs intervals were overlapped. The distribution of different expression LncRNA and the location of QTLs were different in three ecotypes of Brassica napus, which suggesting that LncRNA played an important role in ecotype differentiation and formation of important agronomic traits in Brassica napus.

Key words: Brassica napus, ecotype, long non-coding RNA, flowering genes, QTLs

Fig. 1

Contemporaneous morphology of three ecotypes in Brassica napus L. Morphology of three ecotypes of Brassica napus, about 110 days after sowing in semi-winter environment of Wuhan, in 2021-2022. A: Westar; B: Zhongshuang 11; C: Tapidor. Bar: 20 cm。"

Fig. 2

mRNA-seq analysis of six Brassica napus cultivars in three ecotypes A: mRNA-seq PCA analysis of 12 samples from six Brassica napus cultivars; B: the relative expression level of all genes in six Brassica napus cultivars and three ecotypes; C: the number of differentially expressed genes among three ecotypes; D: Venn diagram of differentially expressed genes among three ecotypes of Brassica napus. SOR: spring type rapeseed; SWOR: semi-winter type rapeseed; WOR: winter-type rapeseed."

Fig. 3

GO function and KEGG pathway enrichment of differentially expressed genes in three ecotypes of Brassica napus A: GO function enrichment of differentially expressed genes in three ecotypes of Brassica napus; B: KEGG pathway enrichment of differentially expressed genes in three ecotypes of Brassica napus."

Fig. 4

LncRNA-seq analysis of six Brassica napus cultivars from three ecotypes A: LncRNA-seq PCA analysis of 12 samples from six Brassica napus cultivars; B: the number of expressed LncRNA in six Brassica napus cultivars and three ecotypes; C: Venn diagram of expressed LncRNA among three ecotypes; D: the distribution of LncRNA on 19 chromosomes and An, Cn subgenomes in three ecotypes of Brassica napus. SOR: spring type rapeseed; SWOR: semi-winter type rapeseed; WOR: winter-type rapeseed."

Fig. 5

GO function and KEGG pathway enrichment of candidate target gene of differentially expressed LncRNA in three ecotypes of Brassica napus A: GO function enrichment of candidate target gene of differentially expressed LncRNA in three ecotypes of Brassica napus; B: KEGG pathway enrichment of candidate target gene of differentially expressed LncRNA in three ecotypes of Brassica napus."

Table S1

Differentially expressed genes related flowering time"

拟南芥基因
Arabidopsis thaliana gene-ID
基因缩写
Symbol
甘蓝型油菜基因ID
Brassica napus gene-ID
开花调控途径
Flowering regulation pathway
AT4G11880 AGL14 A09p27210.1_BnaDAR 其他途径 Other
AT5G13790 AGL15 C09p63310.1_BnaDAR 开花整合基因 Floral integrator
AT1G69120 AP1 A02p15620.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT1G69120 AP1 C02p17480.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT3G21320 AT3G21320 A03p42190.1_BnaDAR 其他途径 Other
AT5G37780 CAM1 C09p30310.1_BnaDAR 其他途径 Other
AT4G17640 CKB2 C01p11310.1_BnaDAR 光信号途径 Light singnalling
AT2G42530 COR15b A03p23680.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b A03p23700.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b C03p28240.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b C03p28260.1_BnaDAR 春化途径 Vernalization
AT1G12610 DDF1 A06p09040.1_BnaDAR 赤霉素途径 Gibberellin
AT1G12610 DDF1 A08p32730.1_BnaDAR 赤霉素途径 Gibberellin
AT1G12610 DDF1 C05p10640.1_BnaDAR 赤霉素途径 Gibberellin
AT1G12610 DDF1 C08p15540.1_BnaDAR 赤霉素途径 Gibberellin
AT1G12610 DDF1 C08p47250.1_BnaDAR 赤霉素途径 Gibberellin
AT5G54510 DFL1 A02p10230.1_BnaDAR 其他途径 Other
AT4G14690 ELIP2 A01p22980.1_BnaDAR 光信号途径 Light singnalling
拟南芥基因
Arabidopsis thaliana gene-ID
基因缩写
Symbol
甘蓝型油菜基因ID
Brassica napus gene-ID
开花调控途径
Flowering regulation pathway
AT4G14690 ELIP2 C01p28420.1_BnaDAR 光信号途径 Light singnalling
AT5G10140 FLC A02p00340.1_BnaDAR 春化途径 Vernalization
AT5G10140 FLC A03p16730.1_BnaDAR 春化途径 Vernalization
AT5G10140 FLC C03p04920.1_BnaDAR 春化途径 Vernalization
AT5G10140 FLC C09p67350.1_BnaDAR 春化途径 Vernalization
AT5G10140 FLC C09p67380.1_BnaDAR 春化途径 Vernalization
AT1G30040 GA2OX2 C03p76330.1_BnaDAR 赤霉素途径 Gibberellin
AT5G67100 ICU2 A02p33280.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT5G15970 KIN2 A02p03320.1_BnaDAR 春化途径 Vernalization
AT5G15970 KIN2 C02p08090.1_BnaDAR 春化途径 Vernalization
AT4G32040 KNAT5 C01p06890.1_BnaDAR 赤霉素途径 Gibberellin
AT5G65060 MAF3 A02p43650.1_BnaDAR 春化途径 Vernalization
AT5G65060 MAF3 C02p63700.1_BnaDAR 春化途径 Vernalization
AT3G01460 MBD9 C09p75140.1_BnaDAR 光周期和生物钟途径Photoperiod and circadian clock
AT3G62090 PIL2 C08p37050.1_BnaDAR 光信号途径 Light singnalling
AT1G13260 RAV1 A09p63190.1_BnaDAR 光周期和生物钟途径Photoperiod and circadian clock
AT2G03710 SEPALLATA4 A02p33530.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT2G45660 SOC1 A05p05620.1_BnaDAR 开花整合基因 Floral integrator
AT2G45660 SOC1 C03p30160.1_BnaDAR 开花整合基因 Floral integrator
AT2G33810 SPL3 C04p15220.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT3G16640 TCTP C01p47690.1_BnaDAR 花分化和发育途径 Floral meristems and development

Table S2

Differentially expressed LncRNA related flowering time gene"

拟南芥基因
Arabidopsis thaliana gene-ID
基因缩写
Symbol
甘蓝型油菜基因ID
Brassica napus gene-ID
开花调控途径
Flowering regulation pathway
AT4G14690 ELIP2 A01p22980.1_BnaDAR 光信号途径 Light singnalling
AT5G15970 KIN2 A02p03320.1_BnaDAR 春化途径 Vernalization
AT1G69120 AP1 A02p15620.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT1G78440 GA2OX1 A02p22190.1_BnaDAR 赤霉素途径 Gibberellin
AT2G42530 COR15b A03p23680.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b A03p23700.1_BnaDAR 春化途径 Vernalization
AT2G45660 SOC1 A03p25280.1_BnaDAR 开花整合基因 Floral integrator
AT2G45660 SOC1 A05p05620.1_BnaDAR 开花整合基因 Floral integrator
AT5G11260 HY5 A10p26460.1_BnaDAR 光信号途径 Light singnalling
AT5G15970 KIN2 C02p08090.1_BnaDAR 春化途径 Vernalization
AT1G69120 AP1 C02p17480.1_BnaDAR 花分化和发育途径 Floral meristems and development
AT4G25470 CBF2 C03p19680.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b C03p28240.1_BnaDAR 春化途径 Vernalization
AT2G42530 COR15b C03p28260.1_BnaDAR 春化途径 Vernalization
AT1G12610 DDF1 C08p47250.1_BnaDAR 赤霉素途径 Gibberellin
AT5G20730 NPH4 C09p55450.1_BnaDAR 光信号途径 Light singnalling

Table S3

Differentially expressed genes and differentially expressed candidate target genes of LncRNA"

基因缩写
Symbol
拟南芥基因
Arabidopsis thaliana gene
甘蓝型油菜基因
Brassica napus gene
非编码RNA
LncRNA
COR15b AT2G42530 A03p23680.1_BnaDAR BnaLncA02_026, BnaLncA02_041, BnaLncA03_157, BnaLncC02_051, BnaLncC03_115, BnaLncC03_185, BnaLncC03_268, BnaLncC05_059, BnaLncC05_060, BnaLncC06_126, BnaLncC06_146
COR15b AT2G42530 A03p23700.1_BnaDAR BnaLncA02_026, BnaLncA02_041, BnaLncA03_157, BnaLncC02_051, BnaLncC03_115, BnaLncC03_185, BnaLncC03_268, BnaLncC05_059, BnaLncC05_060, BnaLncC06_126, BnaLncC06_146
COR15b AT2G42530 C03p28240.1_BnaDAR BnaLncA02_026, BnaLncA02_041, BnaLncA03_109, BnaLncA03_157, BnaLncA03_168, BnaLncC02_040, BnaLncC02_051, BnaLncC03_115, BnaLncC03_185, BnaLncC03_268, BnaLncC05_059, BnaLncC05_060, BnaLncC06_126, BnaLncC06_146, BnaLncC07_243
COR15b AT2G42530 C03p28260.1_BnaDAR BnaLncA02_026, BnaLncA02_041, BnaLncA03_109, BnaLncA03_157, BnaLncA03_168, BnaLncC02_040, BnaLncC02_051, BnaLncC03_115, BnaLncC03_164, BnaLncC03_185, BnaLncC03_268, BnaLncC05_059, BnaLncC05_060, BnaLncC06_126, BnaLncC06_146, BnaLncC07_243
AP1 AT1G69120 A02p15620.1_BnaDAR BnaLncA02_044, BnaLncA06_211, BnaLncC08_129
AP1 AT1G69120 C02p17480.1_BnaDAR BnaLncA01_118, BnaLncA08_034, BnaLncA08_093, BnaLncA10_078, BnaLncC01_179, BnaLncC03_247, BnaLncC04_029
KIN2 AT5G15970 A02p03320.1_BnaDAR BnaLncA02_027, BnaLncA02_142, BnaLncA03_109, BnaLncA03_168, BnaLncA06_218, BnaLncA09_045, BnaLncA09_161, BnaLncC02_040, BnaLncC03_164, BnaLncC05_060, BnaLncC06_146, BnaLncC07_243, BnaLncC08_166
KIN2 AT5G15970 C02p08090.1_BnaDAR BnaLncA02_027, BnaLncA02_142, BnaLncA03_109, BnaLncA03_168, BnaLncA06_218, BnaLncA09_045, BnaLncA09_161, BnaLncC02_016, BnaLncC02_040, BnaLncC03_164, BnaLncC05_060, BnaLncC07_243, BnaLncC08_166
DDF1 AT1G12610 C08p47250.1_BnaDAR BnaLncC08_081
ELIP2 AT4G14690 A01p22980.1_BnaDAR BnaLncA02_026, BnaLncA02_041, BnaLncA03_109, BnaLncA03_157, BnaLncA03_168, BnaLncC02_040, BnaLncC02_051, BnaLncC03_115, BnaLncC03_164, BnaLncC03_185, BnaLncC03_268, BnaLncC05_059, BnaLncC05_060, BnaLncC06_126, BnaLncC06_146, BnaLncC07_243
SOC1 AT2G45660 A05p05620.1_BnaDAR BnaLncA02_027, BnaLncA03_109, BnaLncA03_168, BnaLncA06_218, BnaLncA09_161, BnaLncC02_040, BnaLncC08_166

Fig. 6

Differentially expressed mRNA-LncRNA interaction module and gene expression in population of Brassica napus A: the differentially expressed mRNA-LncRNA interaction module, including 8 genes and 23 LncRNA sequences; B: the relative expression level of COR15b, KIN2, and SOC1 genes (multiple copies) in different ecotypes of Brassica napus population. FPKM: the fragments per kilobase of exon model per million mapped fragments; * indicates significant differences at the 0.05 probability level; ** indicates significant differences at the 0.01 probability level; ns: no significant difference (P > 0.05). SOR: spring type rapeseed; SWOR: semi-winter type rapeseed; WOR: winter-type rapeseed."

Table S4

Summary of collected QTLs in Brassica napus"

QTL性状 QTL trait 数目Number
开花时间Flowering times 181
千粒重Thousand seed weight 160
株高Plant height 124
种子产量Seed yield 89
成熟时间Maturity time 73
第一分支数Number of primary branches 53
其他Others 292
A基因组数目A genome numbers 624
C基因组数目C genome numbers 348
总计Total 972

Fig. 7

Collinearity of agronomic QTLs and identified LncRNA in Brassica napus L."

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