运用小麦杂交株系挖掘环式光合电子传递调控基因并应用于作物高光效改造

doi:10.13560/j.cnki.biotech.bull.1985.2025-0620

摘要/Abstract

摘要：

目的在全球气候变暖以及粮食需求不断增长的背景下，粮食安全问题愈发严峻，要求作物在增产的同时提高环境胁迫适应能力。光合作用是作物产量形成的基础，其光反应阶段的环式光合电子传递途径偶联ATP合成并调节还原力积累，对植物的高温响应和高光效具有重要作用，因此本研究致力于相关遗传调控因子的发掘和应用。方法利用具有环式光合电子传递活性差异的代表性小麦品系为参照，结合光合参数测量、蛋白含量测定和转录组测序技术，从小麦小偃54和京411杂交培育的重组自交系后代中筛选获得了具有环式电子传递活性两极分离的株系及其差异表达基因。结果其中环式电子传递活性较高的株系同时表现出较高的线性电子传递活性和光合CO₂同化速率，并在光强升高条件下保持优势，可作为高光效育种材料加以应用。通过差异表达基因分析挖掘出一系列具有促进环式光合电子传递或整体光合活性潜力的功能基因和转录因子。将部分基因（包括TaPnsL2和TaNAC等）构建过量表达载体转化水稻栽培品种秀水134，T₁代和T₂代转基因材料在海南省和上海大田试验表现出光合速率升高的优势。结论针对环式电子传递活性的遗传筛选或改造，具有提高作物光合效率或强光适应性的潜力。

关键词: 环式电子传递, 光合作用, NDH复合体, 遗传调控, 小麦, 水稻

Abstract:

Objective Global warming and a rapidly increasing global population present formidable challenges to food security, necessitating significant advancements in both crop yield improvement and enhanced resilience to environmental stresses. Photosynthesis serves as the foundation of crop productivity, with the cyclic electron flow during the light reactions coupling ATP synthesis and modulating reducing power accumulation. This pathway plays an important role in heat adaptation and photosynthetic efficiency, making the identification and application of its genetic regulators imperative. Method This study utilized representative wheat lines with varying cyclic photosynthetic electron transport activities as references, combined with photosynthetic parameter measurements, protein content assays, and transcriptome sequencing techniques. From the recombinant inbred lines derived from the crossbreeding of wheat varieties Xiaoyan 54 and Jing 411, strains exhibiting polar separation in cyclic electron transport activity were screened and differentially expressed genes were obtained. Result The strains with higher cyclic electron transport activity also demonstrated elevated linear electron transport activity and photosynthetic CO₂ assimilation rates, maintaining these advantages under increased light intensity, making them suitable for application as high-efficiency breeding materials. Through differential gene expression analysis, a series of functional genes and transcription factors with potential to enhance cyclic photosynthetic electron transport or overall photosynthetic activity were identified. Selected genes (including TaPnsL2 and TaNAC) were constructed into overexpressing vectors and transformed into the rice cultivar Xiushui 134. The T₁ and T₂ generation transgenic materials presented advantages in photosynthetic rate during field trials in both Hainan province and Shanghai. Conclusion Through genetic screening or modification targeting the activity of cyclic electron transport, there is potential to enhance the photosynthetic efficiency or adaptability to high light intensity in crops.

Key words: cyclic electron transport, photosynthesis, NDH complex, genetic regulation, wheat, rice

范艳飞, 叶露幻, 李雨桐, 王钏跞, 张瑞, 罗建华, 王鹏. 运用小麦杂交株系挖掘环式光合电子传递调控基因并应用于作物高光效改造[J]. 生物技术通报, 2025, 41(10): 72-86.

FAN Yan-fei, YE Lu-huan, LI Yu-tong, WANG Chuan-luo, ZHANG Rui, LUO Jian-hua, WANG Peng. Utilizing Wheat Hybrid Lines to Mine Genes Regulating Cyclic Electron Flow and Applying Them in Improving Photosynthetic Efficiency in Crops[J]. Biotechnology Bulletin, 2025, 41(10): 72-86.

图/表 8

图1 小麦重组自交系群体种植及代表性品种的P700+暗还原初始速率和ETR（Ⅰ）-ETR（Ⅱ）测量A：上方为进行第一轮P700+还原速率测定时其中1批的小麦材料实拍图；左下为5个小麦代表性品种；右下为在大田生长的小麦群体。B：左侧为5个小麦代表性品种P700+的还原曲线实测图，右侧为对应的曲线下降初始斜率柱状图。C： 5个小麦代表性品种在光强梯度升高情况下的ETR（Ⅰ）-ETR（Ⅱ）折线图

Fig. 1 Planting of wheat recombinant inbred population, and measurements of initial P700+ dark-reduction rates and ETR(Ⅰ)- ETR(Ⅱ) in representative varietiesA: The above is a photo of one batch of wheat materials during the first round of P700+ reduction rate measurement; the bottom left shows five representative wheat varieties; the bottom right displays the wheat population growing in the field. B: Left, P700+reduction curves of five representative wheat varieties; right, initial slope of curve decline (bar chart). C: Line chart of ETR(Ⅰ)-ETR(Ⅱ) for five representative wheat varieties under increasing light intensity gradients

图2 第一轮测定小麦群体P700+暗还原初始速率分布示例第一轮P700+暗还原初始速率筛选覆盖了整个小麦重组自交系群体，此处显示前80个株系（RL01-RL80，“RL”代表重组自交系）的测量结果，其中每连续的20个株系加上5个代表性品种作为一组进行测量和作图。代表性品种以蓝色显示，在数据分布的两端选出环式电子传递活性高和低的株系，分别以绿色和浅棕色显示

Fig. 2 Distribution examples of initial P700+ dark-reduction rates of wheat population in the first-round measurementThe first-round screening of initial P700+ dark-reduction rates covered the entire wheat recombinant inbred line population. Here, the measurements of the first 80 lines (RL01-RL80, “RL” is short for “recombinant inbred line”) are displayed, with every consecutive 20 lines plus 5 representative cultivars measured and plotted as a group. The representative varieties are displayed in blue, while the lines with high and low cyclic electron transport activity selected from both ends of the data distribution are shown in green and light brown, respectively

图3 第二轮测定小麦群体P700+暗还原初始速率分布图第二轮P700+暗还原初始速率筛选面向第一轮筛选出的环式电子传递活性高和低的株系，将其随机编入4个包含代表性品种的批次采用与第一轮相同的方式进行测量和作图。代表性品种仍以蓝色显示，第一轮测定筛选出的环式电子传递活性高和低的株系，仍以绿色和浅棕色显示。“RL”代表重组自交系

Fig. 3 Distribution of initial P700+ dark reduction rates of wheat population in the second-round measurementThe second-round initial P700+ dark-reduction rate screening focused on lines with either high or low cyclic electron transport activity from the. They were randomly grouped into 4 batches alongside representative cultivars and measured and graphed usingthe same protocol as the first-round screening. Representative varieties are still shown in blue, while the lines with high and low cyclic electron transport activity selected from the first-round screening remain displayed in green and light brown, respectively. “RL” is short for “recombinant inbred line”

图4 代表性小麦品种的生长表型及在梯度光强下的光合参数A：抽穗后的5个代表性小麦品种；B-D：代表性品种在光强梯度下的光合速率Anet、Y（Ⅱ）和ETR（Ⅱ）。大田小麦抽穗后，在晴朗的上午使用Li6400光合仪和PAM2000便携式叶绿素荧光仪测定了小麦代表性品种LX99、SSDL10、XY101、XY54和J411的旗叶在不同光强下的光合速率及电子传递活性，生物学重复为4-6个

Fig. 4 Growth phenotypes of representative wheat cultivars and light-intensity-dependent photosynthetic parametersA: Five representative wheat varieties after heading. B-D: Net photosynthetic rate (Anet), Y(Ⅱ) and ETR(Ⅱ) of representative varieties under light-intensity gradients. Field measurements of photosynthetic rates and electron transport activities under varying light intensities were conducted on flag leaves of wheat representative cultivars (LX99,SSDL10,XY101, XY54, and J411) using Li-6400 photosynthesis system and PAM-2000 portable chlorophyll fluorometer. The measurements were performed on clear mornings during the post-heading stage, with 4-6 biological replicates per cultivar

图5 小麦重组自交株系的生长表型及在梯度光强下的光合参数A：抽穗后的4个小麦重组自交株系；B-D：分别为4个小麦重组自交株系在光强梯度下的光合速率Anet、Y（Ⅱ）和ETR（Ⅱ）。大田小麦抽穗后，在晴朗的上午使用Li6400光合仪和PAM2000便携式叶绿素荧光仪测定了小麦重组自交系群体株系RL18、RL48、RL42和RL71的旗叶在不同光强下的光合速率及电子传递活性，生物学重复为5-6个

Fig. 5 Growth phenotypes of wheat recombinant inbred lines and light-intensity-dependent photosynthetic parametersA: Four wheat recombinant inbred lines after heading. B-D: Net photosynthetic rate (Anet), Y(Ⅱ) and ETR(Ⅱ) of wheat recombinant inbred lines under light-intensity gradients, respectively. Field measurements of photosynthetic rates and electron transport activities under varying light intensities were conducted on flag leaves of wheat recombinant inbred lines (RL18, RL48, RL42 and RL71) using Li-6400 photosynthesis system and PAM-2000 portable chlorophyll fluorometer. The measurements were performed on clear mornings during the post-heading stage, with 5-6 biological replicates per cultivar

图6 重组自交系群体NDH亚基含量测定A：重组自交系群体NDH-H亚基含量测定（第一轮）；B：重组自交系群体NDH含量第二轮测定。每张免疫杂交（Western blot）图内部具有可比性，杂交图之间显色强度可能不同。每张杂交图均含有XY101（NDH-H含量高）和LX99（NDH-H含量低）作为参考

Fig. 6 Determination of NDH subunit content in recombinant inbred line populationA: Determination of NDH-H subunit content in recombinant inbred line population (first round). B: Second-round quantification of NDH complex content in the recombinant inbred line population. The Western blot results are comparable within each blot, but the signal intensities may vary between different blots. Each immunoblotting image contains XY101 (high NDH-H content) and LX99 (low NDH-H content) as references

图7 转录组测序结果分析示例A-D：分别显示光系统间电子递体、环式光合电子传递、光系统I和转录因子相关基因差异表达。热图显示各品种FPKM数值相对于“LX99+J411”平均值的倍数的对数值|Log2（FC）|

Fig. 7 Analysis example of transcriptome sequencing resultsA-D: Differential expressions of genes related to intersystem electron carriers, cyclic photosynthetic electron transport, PSI and transcription factors respectively. The heatmap demonstrates the logarithmic values |Log2（FC）| of the fold change of FPKM values for each variety relative to the average of “LX99+J411”

图8 小麦PnsL2和NAC基因过量表达的T1和T2代水稻株系光合速率分析A-C：小麦PnsL2基因过量表达水稻于海南和上海、不同测量光强下的净光合速率（Anet）；D-F：小麦转录因子基因NAC过量表达水稻于海南和上海、不同测量光强下的净光合速率（Anet）

Fig. 8 Photosynthetic rate analysis of T1 and T2 generation rice lines overexpressing wheat PnsL2 and NAC genesA-C: Net photosynthetic rate (Anet) of wheat PnsL2 gene-overexpressing rice lines under different measuring light intensities in Hainan and Shanghai. D-F: Net photosynthetic rate (Anet) of wheat transcription factor gene NAC-overexpressing rice lines under different measuring light intensities in Hainan and Shanghai

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