NaCl胁迫对不同耐盐性粳稻种质幼苗叶绿素荧光特性的影响

【目的】叶绿素荧光可反映盐胁迫下植物光合机构防御机制的受害程度和抗逆性。通过分析盐胁迫对不同耐盐性粳稻种质叶绿素荧光特性的影响,揭示其诱导动力学特征,初步阐明叶绿素荧光相关基因调控粳稻种质苗期耐盐性的机制,为耐盐水稻品种筛选和培育提供理论依据。【方法】以8份耐盐、8份盐敏感粳稻种质为试验材料,于苗期在水培条件下分别测定两组种质0 mmol·L^-1和125 mmol·L^-1 NaCl胁迫3 d、6 d的叶片叶绿素荧光参数。通过主成分分析筛选关键指标,利用隶属函数和标准差系数赋予权重法综合评价各种质资源耐盐性,获得典型耐（敏）盐种质开展叶绿素荧光相关基因OsHCF222和OsABCI7的表达特异性分析。【结果】与对照（CK,0 mmol·L^-1 NaCl 3 d、6 d）相比,盐胁迫（125 mmol·L^-1 NaCl 3 d、6 d）可显著降低粳稻种质的最大荧光产量（F_m）、原初光能转化效率（F_v/F_m）。与CK相比,耐盐种质的非光化学荧光淬灭系数（NPQ）和可变荧光的非光化学猝灭系数（qN）在盐胁迫3 d时显著降低,初始荧光产量（F_o）在盐胁迫6 d显著升高;盐敏感种质的光化学淬灭系数（qP）和qN在盐胁迫3 d时显著降低,实际光化学量子产量（Y）、NPQ、表观光合电子传递速率（ETR）在盐胁迫3 d、6 d时极显著降低。盐胁迫下F_m、F_v/F_m、Y、NPQ和ETR与耐盐级别（STS）均呈极显著正相关,且在耐、敏盐种质间差异显著。通过主成分分析将8个叶绿素荧光参数转换为2个主成分,累积贡献率88.018%;结合各因子载荷大小筛选出F_m、F_v/F_m、Y、NPQ和ETR 5个关键指标,可将16份粳稻种质明确划分为耐盐组和盐敏感组两类。以两个主成分的隶属函数结合权重处理并累加计算盐胁迫下叶绿素荧光特性综合评价值D（D_CF）,并依此获得16份粳稻种质排名。采用Kinetic模式测定盐胁迫和正常生长（CK）条件下耐盐种质Cigalon、Bertone和盐敏感种质新竹8号、幸实的叶绿素荧光诱导动力学曲线。在CK条件下,4份粳稻种质表现为相似的曲线形状,斜率较大,P峰出现时间基本相同;盐胁迫下,随着盐胁迫时间的延长,盐敏感种质的P峰迅速降低,M峰和曲线斜率逐渐变小;耐盐种质仍维持较高的P峰,M峰和曲线斜率与CK相比无明显变化。通过对综合排名第1的耐盐种质Cigalon和排名第16的盐敏感种质幸实NaCl胁迫不同时间叶绿素荧光相关基因OsHCF222和OsABCI7的qPCR分析,结合叶绿素含量、耐盐相关叶绿素荧光参数的动态变化和相关分析,初步明确了OsHCF222和OsABCI7的差异表达与粳稻种质苗期耐盐性的关系。【结论】不同耐盐性粳稻种质的叶绿素荧光参数对盐胁迫的响应不尽相同,F_m、F_v/F_m、Y、NPQ和ETR与水稻苗期耐盐性密切相关;OsHCF222和OsABCI7的表达量直接影响了粳稻种质苗期耐盐性;在耐盐粳稻中,NPQ和F_v/F_m起关键作用,F_m可能在调节盐敏感粳稻耐盐性中发挥重要作用。

Effects of NaCl Stress on the Chlorophyll Fluorescence Characteristics of Seedlings of Japonica Rice Germplasm with Different Salt Tolerances

【Objective】Chlorophyll fluorescence parameters can reflect the damage degrees and stress resistance of the plant photosynthetic machinery under salt stress. In this study, the analysis of effects of salt stress on chlorophyll fluorescence characteristics of japonica rice with different salt tolerances was performed to reveal its induced kinetic characteristics and preliminarily elucidate the mechanism of OsHCF222 and OsABCI7 regulating salt tolerance of japonica rice at seedling stage, so as to provide a theoretical foundation for screening and breeding salt-tolerant rice varieties.【Method】Eight salt-tolerant and eight salt-sensitive japonica rice germplasm accessions were used as experimental materials in this study. Leaf chlorophyll fluorescence parameters were measured for these materials after treatment using 0 mmol·L^-1 or 125 mmol·L^-1 NaCl for 3 or 6 days. Principal component analysis (PCA) was used to screen the key indexes for salt tolerance, and a comprehensive evaluation of japonica rice germplasm was carried out with membership functions and weighted standard deviation coefficient method. The resulting salt-tolerant and salt-sensitive germplasm were used to analyze the relative expressions of OsHCF222 and OsABCI7, two chlorophyll fluorescence related genes. 【Result】Compared with the control (CK, 0 mmol·L^-1 NaCl for 3 days or 6 days), the salt stress treatment (125 mmol·L^-1 NaCl 3 days or 6 days) significantly reduced the maximal fluorescence (F_m) and maximum quantum efficiency of PSII (F_v/F_m) of japonica rice germplasm. For salt tolerant germplasm accessions, the non-photochemical quenching coefficient (NPQ) and coefficient of non-photochemical quenching of variable fluorescence (qN) decreased significantly at 3 days after salt stress, while the initial fluorescence yield (F_o) increased significantly at 6 days after salt stress. The photochemical quenching coefficient (qP) and qN of the salt-sensitive germplasm accessions decreased significantly on the 3rd day after salt stress treatment, while the indexes including yield (Y), NPQ and photosynthetic electron transfer rate (ETR) decreased significantly on the 3rd and 6th day after salt stress treatment. Under salt stress, F_m, F_v/F_m, Y, NPQ and ETR were positively correlated with salt tolerance score (STS), and there were significant differences between salt-tolerant and salt-sensitive japonica rice germplasm accessions. PCA with eight chlorophyll fluorescence parameters revealed two major components, with a cumulative contribution rate of 88.018%. Five key indexes, including F_m, F_v/F_m, Y, NPQ and ETR, were selected based on the loading of each component. The 16 accessions were subsequently assigned to salt tolerant and salt sensitive groups by cluster analysis. A comprehensive evaluation value D (D_CF) of chlorophyll fluorescence characteristics under salt stress was obtained by using membership function combined with the index weight method, and then the ranking of the 16 accessions was obtained. Chlorophyll fluorescence induction curves of salt-tolerant Cigalon and Bertone, salt-sensitive Xinzhu8 and Sachiminori under salt stress and CK were created using the Kinetic model. Under CK condition, the four japonica rice germplasm accessions showed similar curve shapes with a large slope and the occurrence time of P peak was basically the same. Under the salt stress treatment, peak P, peak M and the curve slope of salt-sensitive accessions decreased rapidly, while the salt-tolerant accessions still maintained high P peak and curve slope. Through quantitative real-time PCR analysis of OsHCF222 and OsABCI7 in Cigalon and Sachiminori at different times under NaCl stress, the dynamic changes and correlation between leaf chlorophyll content and five key chlorophyll fluorescence parameters, the potential role of the two genes in salt tolerance of japonica rice were preliminarily clarified. 【Conclusion】Chlorophyll fluorescence parameters of japonica rice germplasm with different salt tolerances responded differently to salt stress. F_m, F_v/F_m, Y, NPQ and ETR were closely related to salt tolerance in rice. The expression levels of OsHCF222 and OsABCI7 directly affected the salt tolerance of japonica rice germplasm at seedling stage. In salt-tolerant japonica rice, NPQ and F_v/F_m played as key indexes, while F_m might play an important role in salt-sensitive japonica rice.