水稻叶和籽粒镉积累机制及QTL定位研究

doi:10.13560/j.cnki.biotech.bull.1985.2021-1192

摘要/Abstract

摘要：

水稻是我国最重要的粮食作物,挖掘控制水稻叶和籽粒镉积累的关键基因,阐明水稻镉积累和转运的遗传机制,为培育籽粒低镉积累的水稻品种、保障粮食安全和人类健康提供依据。通过离子组学技术系统分析中国栽培稻核心种质资源209个品种籽粒的离子谱,鉴定出一批铁、锌等高富集以及镉低积累的水稻品种,并选择籼稻（indica）品种花楸03和粳稻（japonica）品种SKC进行进一步研究。结果表明,尽管这两个品种对镉的耐受性和吸收能力无显著差异,花楸03籽粒和地上部分的镉积累量显著高于SKC,而铁、锌等含量差异不显著。以花楸03和SKC为亲本构建了包含137个单株的双单倍体（doubled haploid,DH）群体,共检测到8个控制镉在叶和籽粒中积累的QTL,分别位于第2、3、4、7、8、10和11染色体上,能解释10.6%-39.4%的表型变异。其中第3染色体上检测到的控制镉向籽粒转运的QTL qGCd3定位在RM6266-RM2334,LOD（limit of detection）值和贡献率分别为3.81和39.4%,此处可能存在一个控制镉向籽粒转运的重要基因。

关键词: 水稻, 核心种质, 镉, QTL定位

Abstract:

Rice is the most important staple food in China,thus mining the key genes controlling cadmium（Cd）accumulation in rice leaves and grains and clarifying the genetic mechanism of Cd accumulation and transport may provide the basis for breeding rice varieties with low cadmium accumulation and ensuring food security and human health. Inomics screening was performed to systemically profile ionic spectrum in the grains of 209 rice varieties from the Rice Core Germplasm cultivated in China. The accessions with differential cadmium and/or iron（Fe）/zinc（Zn）accumulations were identified,from which indicia cultivar HQ03 and japonica cultivar SKC were selected for further study. Results showed that there was no significant difference in cadmium tolerance and uptake between the two varieties,and Cd accumulations in the HQ03 shoots and grains were significantly higher than those in SKC,while no difference in Fe,Zn,etc. Using a doubled haploid（DH）population derived from HQ03 and SKC of 137 individuals,8 quantitative trait loci（QTL）controlling Cd accumulation in the leaves and grains were identified on chromosome 2,3,4,7,8,10 and 11,accounting for 10.6%-39.4% of the phenotypic variance detected. Among them,qGCd3,controlling Cd transport to grains,was delimited in the interval of RM6266-RM2334 on chromosome 3,with an LOD（limit of detection）value of 3.81 and a phenotypic contribution of 39.4%,indicating that an important gene controlling cadmium transport to grain may exist here.

Key words: rice, core germplasm, cadmium, QTL mapping

黄婧, 朱亮, 薛蓬勃, 付强. 水稻叶和籽粒镉积累机制及QTL定位研究[J]. 生物技术通报, 2022, 38(8): 118-126.

HUANG Jing, ZHU Liang, XUE Peng-bo, FU Qiang. Research on Mechanism and QTL Mapping Associated with Cadmium Accumulation in Rice Leaves and Grains[J]. Biotechnology Bulletin, 2022, 38(8): 118-126.

图/表 11

图1 核心种质资源水稻籽粒中Cd的积累 Z值=（样本元素浓度-平均值）/标准差,用于表达样品中某一元素偏离群体平均值的程度。X轴表示水稻品种的编号

Fig. 1 Profiling of Cd accumulation in grains of rice core germplasm Z value =（individual value-population average）/population SD,and represents SD from the average value of the whole population. X axis refers to the serial number of rice cultivars

图2 水稻籽粒中Cd2+、Fe2+、Zn2+和Mn2+元素的相关关系

Fig. 2 Correlation among Cd2+, Fe2+, Zn2+ and Mn2+ elements in rice grains

表1 大田、镉污染水槽和对照水槽土壤中Cd等元素的含量

Table 1 Contents of Cd and other metals in the soil of contaminated paddy field,contaminated sink and control sink （μg·g-1 DW）

土壤类型Soil type	镉Cd	铁Fe	锌Zn	铜Cu	锰Mn	砷As
对照水槽Control sink	0.14±0.02	19 047.64±232.07	157.34±8.26	23.51±9.55	318.16±3.63	9.69±2.69
污染水槽 Contaminated sink	1.09±0.11	20 495.83±270.84	271.98±9.51	45.39±8.31	322.02±9.58	12.99±1.28
污染大田土壤Contaminated paddy field	0.55±0.07	19 603.91±267.35	265.89±4.15	48.20±3.93	330.8±14.11	12.53±1.34

图3 花楸03与SKC的籽粒和精米中的Cd含量 A：分别种植在污染大田、污染水槽和对照水槽中,花楸03与SKC籽粒中的Cd含量;B：种植在镉污染水槽中,花楸03与SKC精米中的Cd含量。**：P<0.01。下同

Fig.3 Cd accumulation in the grains and milled rice of HQ03 and SKC A：Cd accumulation in the grains of HQ03 and SKC from the rice grown in the contaminated paddy field,contaminated and control sink,respectively. B：Cd accumulation in the milled rice of HQ03 and SKC from the rice grown in the contaminated sink. **：P < 0.01. The same below

图4 花楸03与SKC叶片中的元素积累 A：花楸03和SKC叶中的Cd含量;B、C：花楸03和SKC叶中的Fe等金属元素的含量（B）和K等大量元素的含量（C）

Fig.4 Elements accumulation in the leaves of HQ03 and SKC A：Cd accumulation in the leaves of HQ03 and SKC. B, C：Contents of metal elements such Fe（B）and major elements such as K（C）in the leaves of HQ03 and SKC

图5 不同浓度和不同时间CdCl2处理后花楸03和SKC幼苗叶片与根中Cd的积累 A：不同浓度CdCl2处理7 d后,叶片中Cd的含量;B：用10 μmol/L CdCl2处理不同时间后,叶片中Cd的含量;C：不同浓度CdCl2处理7 d后,根中Cd的含量;D：用10 μmol/L CdCl2处理不同时间后,根中Cd的含量

Fig.5 Cd accumulation in the leaves and roots of HQ03 and SKC from rice seedlings exposed to CdCl2 for indicated concentrations and days A：Cd levels in leaves from rice seedlings exposed to CdCl2 for 7 d at indicated concentrations;B：Cd levels in leaves from rice seedlings exposed to 10 μmol/L CdCl2 for indicated days;C：Cd levels in roots from rice seedlings exposed to CdCl2 for 7 d at indicated concentrations;D：Cd levels in roots from rice seedlings exposed to 10 μmol/L CdCl2 for indicated days

图6 花楸03与SKC对Cd的耐受性分析 A：不同浓度Cd处理下,花楸03和SKC地上部鲜重（FW）;B：不同浓度Cd处理下,花楸03和SKC地下部鲜重（FW）

Fig. 6 Tolerance assay of HQ03 and SKC to Cd A：Fresh weight（FW）of shoots of HQ03 and SKC under different concentrations of Cd treatment. B：Fresh weight（FW）of roots of HQ03 and SKC under different concentrations of Cd treatment

图7 花楸03和SKC的根吸收试验 A：正常培养条件（28℃）;B：低温条件（4℃）

Fig.7 Root uptake assay of HQ03 and SKC A：Normal culture condition（28℃）. B：Low temperature culture condition（4℃）

表2 水稻叶Cd积累的QTL定位

Table 2 Quantitative trait loci（QTLs）mapping for Cd accumulation in rice leaves

数量性状基因座位QTL	染色体Chr.	标记区间Marker interval	LOD值LOD value	贡献率 R²/%	加性效应Additive effect
qLCd2	2	RM450-RM5472	2.70	13.4	12.792
qLCd4	4	RM307-RM401	2.14	11.1	11.234
qLCd10	10	RM271-RM258	3.02	10.6	-0.081 8
qLCd11	11	RM167-RM220	2.07	11.0	7.576 1

表3 水稻籽粒Cd积累的QTL定位

Table 3 Quantitative trait loci（QTLs）mapping for Cd accumulation in rice grains

数量性状基因座位QTL	染色体Chr.	标记区间Marker interval	LOD值LOD value	贡献率R²/%	加性效应Additive effect
qGCd2	2	RM324-RM341	3.39	20.2	-0.187 4
qGCd3	3	RM6266-RM2334	3.81	39.4	0.125 4
qGCd7	7	RM6574-RM6449	2.51	13.2	-5.381 7
qGCd8	8	RM1235-RM1376	3.15	20.4	-0.278 1

图8 水稻叶和籽粒Cd积累的QTL定位示意图

Fig.8 QTLs mapping of Cd accumulation in the rice leaves and grains

参考文献 23

[1]	Haider FU, Liqun C, Coulter JA, et al. Cadmium toxicity in plants:Impacts and remediation strategies[J]. Ecotoxicol Environ Saf, 2021, 211:111887. doi: 10.1016/j.ecoenv.2020.111887 URL
[2]	Chen HM, Chen WC, Chen YX, et al. Pollution of heavy metals in soil-plant system[M]. Beijing: Science Press, 1996.
[3]	Song Y, Wang Y, Mao WF, et al. Dietary cadmium exposure assessment among the Chinese population[J]. PLoS One, 2017, 12(5):e0177978. doi: 10.1371/journal.pone.0177978 URL
[4]	Nawrot TS, Staessen JA, Roels HA, et al. Cadmium exposure in the population:from health risks to strategies of prevention[J]. Biometals, 2010, 23(5):769-782. doi: 10.1007/s10534-010-9343-z pmid: 20517707
[5]	Meng L, Huang TH, Shi JC, et al. Decreasing cadmium uptake of rice(Oryza sativa L.)in the cadmium-contaminated paddy field through different cultivars coupling with appropriate soil amendments[J]. J Soils Sediments, 2019, 19(4):1788-1798. doi: 10.1007/s11368-018-2186-x URL
[6]	Mendoza-Cózatl DG, Jobe TO, Hauser F, et al. Long-distance transport, vacuolar sequestration, tolerance, and transcriptional responses induced by cadmium and arsenic[J]. Curr Opin Plant Biol, 2011, 14(5):554-562. doi: 10.1016/j.pbi.2011.07.004 pmid: 21820943
[7]	Zhao JL, Yang W, Zhang SH, et al. Genome-wide association study and candidate gene analysis of rice cadmium accumulation in grain in a diverse rice collection[J]. Rice:N Y, 2018, 11(1):61.
[8]	Xue DW, Chen MC, Zhang GP. Mapping of QTLs associated with cadmium tolerance and accumulation during seedling stage in rice(Oryza sativa L.)[J]. Euphytica, 2008, 165(3):587-596. doi: 10.1007/s10681-008-9785-3 URL
[9]	Ueno D, Kono I, Yokosho K, et al. A major quantitative trait locus controlling cadmium translocation in rice(Oryza sativa)[J]. New Phytol, 2009, 182(3):644-653. doi: 10.1111/j.1469-8137.2009.02784.x pmid: 19309445
[10]	Ishikawa S, Abe T, Kuramata M, et al. A major quantitative trait locus for increasing cadmium-specific concentration in rice grain is located on the short arm of chromosome 7[J]. J Exp Bot, 2010, 61(3):923-934. doi: 10.1093/jxb/erp360 pmid: 20022924
[11]	Ueno D, Yamaji N, Kono I, et al. Gene limiting cadmium accumulation in rice[J]. Proc Natl Acad Sci USA, 2010, 107(38):16500-16505. doi: 10.1073/pnas.1005396107 URL
[12]	Miyadate H, Adachi S, Hiraizumi A, et al. OsHMA3, a P1B-type of ATPase affects root-to-shoot cadmium translocation in rice by mediating efflux into vacuoles[J]. New Phytol, 2011, 189(1):190-199. doi: 10.1111/j.1469-8137.2010.03459.x URL
[13]	Huang Y, Sun CX, Min J, et al. Association mapping of quantitative trait loci for mineral element contents in whole grain rice(Oryza sativa L.)[J]. J Agric Food Chem, 2015, 63(50):10885-10892. doi: 10.1021/acs.jafc.5b04932 URL
[14]	Wang FJ, Wang M, Liu ZB, et al. Different responses of low grain-Cd-accumulating and high grain-Cd-accumulating rice cultivars to Cd stress[J]. Plant Physiol Biochem, 2015, 96:261-269. doi: 10.1016/j.plaphy.2015.08.001 URL
[15]	Luo JS, Huang J, Zeng DL, et al. A defensin-like protein drives cadmium efflux and allocation in rice[J]. Nat Commun, 2018, 9(1):645. doi: 10.1038/s41467-018-03088-0 URL
[16]	Hu DW, Sheng ZH, Li QL, et al. Identification of QTLs associated with cadmium concentration in rice grains[J]. J Integr Agric, 2018, 17(7):1563-1573. doi: 10.1016/S2095-3119(17)61847-1 URL
[17]	Liu WQ, Pan XW, Li YC, et al. Identification of QTLs and validation of qCd-2 associated with grain cadmium concentrations in rice[J]. Rice Sci, 2019, 26(1):42-49. doi: 10.1016/j.rsci.2018.12.003 URL
[18]	Yan HL, Xu WX, Xie JY, et al. Variation of a major facilitator superfamily gene contributes to differential cadmium accumulation between rice subspecies[J]. Nat Commun, 2019, 10(1):2562. doi: 10.1038/s41467-019-10544-y URL
[19]	Pan XW, Li YC, Liu WQ, et al. QTL mapping and candidate gene analysis of cadmium accumulation in polished rice by genome-wide association study[J]. Sci Rep, 2020, 10(1):11791. doi: 10.1038/s41598-020-68742-4 URL
[20]	Lincoln SE, Daly MJ, Lander ES. Constructing genetic linkage maps with mapmaker/Exp version 3.0: A tutorial and reference manual[M]. 3rd ed. Cambridge: Whitehead Institute for Biometrical Research, 1993.
[21]	Lincoln SE, Daly MJ, Lander ES. Mapping genes controlling quantitative traits using MAPMAKER/QTL version 1.1: A tutorial and reference manual[M]. 2nd ed. Cambridge: Whitehead Institute for Biometrical Research, 1993.
[22]	Uraguchi S, Mori S, Kuramata M, et al. Root-to-shoot Cd translocation via the xylem is the major process determining shoot and grain cadmium accumulation in rice[J]. J Exp Bot, 2009, 60(9):2677-2688. doi: 10.1093/jxb/erp119 pmid: 19401409
[23]	Takahashi A, Agrawal GK, Yamazaki M, et al. Rice Pti1a negatively regulates RAR1-dependent defense responses[J]. Plant Cell, 2007, 19(9):2940-2951. pmid: 17890377