基于QTL和转录组测序鉴定甘蓝型油菜耐旱候选基因

doi:10.3724/SP.J.1006.2024.34144

Abstract

Abstract:

Drought stress severely limits planting promotion and yield increase in Brassica napus L. Drought tolerance is a complex quantitative trait controlled by multiple genes. Combining QTL and transcriptome is an effective method for identifying candidate genes associated with drought tolerance in B. napus. In this study, the seedlings of F_2:6and F_2:8 recombinant inbred lines, constructed by Sanliu’ai (drought sensitivity line) and Kelina-2 (drought tolerance line), were treated with drought stress and well watering at seedling stage. Shoot fresh weight, shoot dry weight, leaf relative water content, malondialdehyde content, and soluble sugar content were measured. The QTL and candidate intervals were identified based on genetic linkage maps, which were constructed using SSR and SNP markers with polymorphism. Subsequently, candidate genes associated with drought tolerance were screened by combining transcriptome sequencing of No11 (drought tolerance material) and No28 (drought sensitivity material). Drought stress decreased shoot fresh weight, shoot dry weight, and leaf relative water content, and increased the contents of malondialdehyde and soluble sugar. QTL and candidate intervals related to drought tolerance were distributed on chromosome A01, A02, A06, A08, A09, A10, C02, C03, C04, C06, and C09. By transcriptome analysis of drought tolerance and sensitivity materials under well water, drought stress for 24, 36, and 48 h, the major different expression genes were enriched in the pathways associated with photosynthesis, fatty acid metabolism, amino acid metabolism, plant hormone signal transduction, ribosome, circadian rhythm and biosynthesis of keratin, cork and wax. A total of 28 candidate genes related to drought tolerance were identified by combining QTL and transcriptome. They coded FLC, bHLH105, TGA4, TEM1, ERF003, ACO3, CHLI1, LHCB6, PORC, etc., which had transcription factor activity, ethylene production and signal transduction, chlorophyll biosynthesis and binding, chlorophyll oxidoreductase and encoding ribosome proteins. These results could provide a basis for revealing drought tolerance mechanism and molecular breeding of drought tolerance variety in B. napus.

Key words: Brassica napus L., drought tolerance, QTL, transcriptome, candidate genes

LI Yang-Yang, WU Dan, XU Jun-Hong, CHEN Zhuo-Yong, XU Xin-Yuan, XU Jin-Pan, TANG Zhong-Lin, ZHANG Ya-Ru, ZHU Li, YAN Zhuo-Li, ZHOU Qing-Yuan, LI Jia-Na, LIU Lie-Zhao, TANG Zhang-Lin. Identification of candidate genes associated with drought tolerance based on QTL and transcriptome sequencing in Brassica napus L.[J].Acta Agronomica Sinica, 2024, 50(4): 820-835.

Figures/Tables 11

Table 1

Table 2

Fig. 1

Fig. 2

Fig. 3

Fig. 4

Table 3

QTL and candidate range of traits associated with drought tolerance in rapeseed"

群体 Population	QTL	置信区间 Confidence interval (cM)	邻近标记 Marker	物理位置 Physical position (bp)	阈值 LOD	染色体 Chr.	候选区间 Candidate range (bp)
F_2:6	qSFWCK-A01	78.85-86.82	AX-95683221	4227188	3.04	A01	4047188-4407188
			AX-179306831	4399498	2.89	A01	4219498-4579498
F_2:8	qRWCDRI-A02	17.64-31.87	AX-177910459	262823	3.07	A02	82823-442823
F_2:8	qRWCDS-A02	16.64-33.30	AX-177910459	262823	2.99	A02	82823-442823
F_2:8	qSFWCK-A02	43.41-47.22	AX-177829332	4692035	2.53	A02	4512035-4872035
F_2:8	qSUGCK-A02	0.00-8.00	AX-177834286	20489037	3.20	A02	20309037-20669037
F_2:6	qSDWCK-A02	0.00-11.74	SWUA02_198	893419-893684	3.14	A02	713419-1073684
			A02_2	893423-893813	3.47	A02	713423-1073813
F_2:8	qSFWDRI-A06	136.38-140.39	AX-95506320	20609909	2.50	A06	20429909-20789909
F_2:8	qMDACK-A08	28.40-30.85	AX-95638560	11831251	2.58	A08	11331251-12331251
F_2:8	qSFWDRI-A08	89.56-109.10	AX-95637797	16969405	3.04	A08	16469405-17469405
F_2:6	qSDWCK-A09	37.27-44.43	AX-179306992	2292222	2.59	A09	1912222-2672222
F_2:6	qSFWCK-A09	37.27-44.43	AX-179306992	2292222	2.70	A09	1912222-2672222
F_2:8	qMDADRI-A09	163.36-165.01	AX-177830392	26135201	4.09	A09	25755201-26515201
F_2:6	qMDACK-A09	0.00-2.00	nia_m046	21769576-21769946	2.91	A09	21389576-22149946
F_2:6	qRWCDS-A09	0.00-2.00	nia_m046	21769576-21769946	2.87	A09	21389576-22149946
F_2:8	qRWCCK-A10	12.22-14.56	AX-182144456	885133	3.06	A10	675133-1095133
F_2:6	qSFWDRI-A10	134.63-148.54	AX-177911667	16025485	3.04	A10	15815485-16235485
			AX-95637504	16164216	3.08	A10	15954216-16374216
			AX-182169619	16167478	3.04	A10	15957478-16377478
F_2:8	qSUGCK-C02	7.00-45.13	AX-182158861	16526368	2.58	C02	15326368-17726368
			AX-95636960	16803922	3.01	C02	15603922-18003922
			AX-182136799	24120437	2.99	C02	22920437-25320437
			AX-182139036	28872169	3.49	C02	27672169-30072169
			AX-182125192	33288173	2.98	C02	32088173-34488173
			AX-105306912	34988363	2.58	C02	33788363-36188363
			AX-95505280	41712868	2.96	C02	40512868-42912868
F_2:6	qRWCDRI-C02	29.49-31.89	SWUC316	33799496-33799694	3.36	C02	32599496-34999694
F_2:6	qRWCDS-C02	29.49-30.89	SWUC316	33799496-33799694	2.77	C02	32599496-34999694
F_2:6	qMDADS-C02	29.89-37.57	SWUC316	33799496-33799694	3.35	C02	32599496-34999694
			SWUC344	40102848-40103191	2.53	C02	38902848-41303191
			SWUC284	43919221-43919418	2.54	C02	42719221-45119418
			SWUC293	36909045-36909237 19357230-19357429	3.00	C02 A02	35709045-38109237 19177230-19537429
F_2:8	qSFWDRI-C03	153.23-157.21	AX-95636893	9150013	2.71	C03	8750013-9550013
F_2:6	qSUGDRI-C04	103.69-108.02	AX-95506064	7272757	7.10	C04	6682757-7862757
F_2:8	qSUGDS-C06	74.57-101.32	AX-105336545	29172287	3.04	C06	28772287-29572287
			AX-95665032	29186383	2.66	C06	28786383-29586383
			AX-86230901	29308911	3.04	C06	28908911-29708911
			AX-95665273	29401522	3.04	C06	29001522-29801522
			AX-95664947	29433854	3.04	C06	29033854-29833854
			AX-105310203	29489446	3.04	C06	29089446-29889446
			AX-105308711	29627549	3.04	C06	29227549-30027549
			AX-86230806	29710301	3.04	C06	29310301-30110301
			AX-182145054	29797943	3.04	C06	29397943-30197943
			AX-86215830	30202409	3.04	C06	29802409-30602409
			AX-86230835	30204371	2.92	C06	29804371-30604371
			AX-105308457	30304436	3.04	C06	29904436-30704436
			AX-95665820	30314052	3.04	C06	29914052-30714052
			AX-105310125	30791904	3.04	C06	30391904-31191904
			AX-182087795	31059928	3.04	C06	30659928-31459928
			AX-95635741	31183285	3.04	C06	30783285-31583285
			AX-86230832	31264725	3.11	C06	30864725-31664725
			AX-177912717	31340640	2.95	C06	30940640-31740640
F_2:8	qSDWDRI-C09	19.55-37.10	AX-95509234	35782323	2.70	C09	35072323-36492323
			AX-86236963	37494562	2.68	C09	36784562-38204562
			AX-182158558	46574561	2.68	C09	45864561-47284561

Table 3

Fig. 5

Fig. 6

Fig. 7

Table 4

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群体 Population	变异来源 Source of variation	RWC		SFW		SDW		SUG		MDA
群体 Population	变异来源 Source of variation	DF	F-value	DF	F-value	DF	F-value	DF	F-value	DF	F-value
F_2:6	水分间Water	1	11.18^**	1	850.03^**	1	214.32^**	1	30.86^**	1	91.75^**
	家系间Line	103	4.64^**	104	14.41^**	104	14.96^**	104	15.29^**	104	552.39^**
	水分×家系 Water×Line	103	3.10^**	104	10.55^**	104	5.98^**	104	9.96^**	104	392.10^**
	误差Error	208		210		210		210		210
F_2:8	水分间Water	1	3175.43^**	1	2237.27^**	1	389.89^**	1	1490.35^**	1	1139.48^**
	家系间Line	125	2.36^**	125	6.25^**	125	6.21^**	125	15.72^**	125	21.62^**
	水分×家系 Water×Line	125	2.30^**	124	3.65^**	124	1.76^**	124	13.32^**	123	11.86^**
	误差Error	446		426		414		462		471

性状 Trait	群体 Population	处理 Treatment	亲本Parent		群体Population
性状 Trait	群体 Population	处理 Treatment	三六矮 Sanliu’ai	科里纳-2 Kelina-2	平均值 Average	标准误差 SE	变幅 Range	峰度 Kurtosis	偏度 Skewness
RWC (%)	F_2:6	CK	91.72	93.35	86.51	0.54	68.96-96.55	0.95	-0.76
		DS	83.98	86.24	85.16	0.58	61.78-96.87	2.27	-1.19
	F_2:8	CK	88.00	86.84	89.74	0.28	72.10-95.14	7.26	-1.64
		DS	65.46	76.11	71.27	0.42	60.83-82.91	-0.55	-0.06
SFW (g)	F_2:6	CK	20.18	10.78	16.55	0.46	4.79-30.27	0.01	0.21
		DS	15.67	9.00	11.71	0.36	4.79-23.90	0.15	0.71
	F_2:8	CK	44.78	27.06	36.79	1.02	11.84-75.42	1.57	1.04
		DS	19.22	22.70	14.36	0.38	6.58-25.85	-0.03	0.62
SDW (g)	F_2:6	CK	1.27	0.72	1.18	0.04	0.41-2.34	0.06	0.64
		DS	0.95	0.61	0.95	0.03	0.37-1.92	0.47	0.87
	F_2:8	CK	3.80	2.65	2.72	0.08	0.89-5.90	0.85	0.74
		DS	2.30	2.10	1.81	0.05	0.59-3.48	0.90	0.59
SUG (μg g^-1)	F_2:6	CK	1496.20	542.03	2744.17	226.85	186.82-16219.47	12.23	2.83
		DS	6485.20	1081.01	3271.71	249.66	222.49-12447.88	3.50	1.69
	F_2:8	CK	3888.00	4581.34	5349.75	131.73	2144.61-10444.09	0.82	0.75
		DS	6265.26	7643.01	7278.97	131.53	3370.63-10742.93	0.17	0.15
MDA (μmol g^-1)	F_2:6	CK	7.49	13.97	9.80	0.44	3.43-34.67	8.58	2.26
		DS	12.77	17.70	10.04	0.34	5.28-23.09	1.81	1.31
	F_2:8	CK	41.18	20.43	29.26	0.97	9.34-64.77	0.54	0.83
		DS	50.15	50.63	41.03	1.04	23.76-71.73	-0.63	0.51

Identification of candidate genes associated with drought tolerance based on QTL and transcriptome sequencing in Brassica napus L.

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QTL	候选基因Candidate gene	QTL	候选基因Candidate gene
qSFWCK-A01	BnaA01g08750D (MYC4) BnaA01g09270D (CHLI1)	qMDADRI-A09	BnaA09g36380D (RR9)
qRWCDRI-A02	BnaA02g00310D (TGA4)	qMDACK-A09	BnaA09g28670D (TEM1) BnaA09g29410D (PRE6)
qRWCDS-A02	BnaA02g00310D (TGA4)	qRWCDS-A09	BnaA09g28670D (TEM1) BnaA09g29410D (PRE6)
qSFWCK-A02	BnaA02g09540D (RPL24)	qRWCCK-A10	BnaA10g01420D (EL22Z) BnaA10g02050D (PORC)
qSUGCK-A02	BnaA02g27860D (CYP71B22) BnaA02g27940D (bHLH105)	qSFWDRI-A10	BnaA10g25310D (SUVH1)
qSDWCK-A02	BnaA02g01690D (RPL10)	qSUGCK-C02	BnaC02g20870D (HPA1)
qMDACK-A08	BnaA08g13440D (RPL28C) BnaA08g14220D (ACO3)	qMDADS-C02	BnaC02g40790D (KNAT3) BnaC02g40810D (ERF003)
qSFWDRI-A08	BnaA08g23760D (LHCB6) BnaA08g25090D (E2-OGDH) BnaA08g25130D (ADT1)	qSFWDRI-C03	BnaC03g17470D (PRK1) BnaC03g17780D (AIP1) BnaC03g18600D (PXG3)
qSDWCK-A09	BnaA09g04650D (ERF003) BnaA09g05170D (RUP2)	qSDWDRI-C09	BnaC09g45770D (RPL37B) BnaC09g46500D (FLC)
qSFWCK-A09	BnaA09g04650D (ERF003) BnaA09g05170D (RUP2)

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