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作物学报

作物学报 ›› 2025, Vol. 51 ›› Issue (6): 1423-1434.doi: 10.3724/SP.J.1006.2025.44198

• 作物遗传育种·种质资源·分子遗传学 •    下一篇

基于改进U-Net++模型的油菜pol TCMS温敏两系育性等级鉴定及温度育性关系的量化研究

李世鹏1(), 陈才武2, 张晶1, 吕恬1, 傅廷栋1, 易斌1,*()   

  1. 1华中农业大学植物科学技术学院, 湖北武汉430070
    2广东韶关烟叶复烤有限公司, 广东韶关 512023
  • 收稿日期:2024-12-01 接受日期:2025-03-26 出版日期:2025-06-12 网络出版日期:2025-04-10
  • 通讯作者: *易斌, E-mail: yibin@mail.hzau.edu.cn
  • 作者简介:E-mail: lisp@webmail.hzau.edu.cn
  • 基金资助:
    本研究由国家重点研发计划项目(2021YFD1600500);武汉市生物育种关键技术攻关及新品种培育科技重大专项(2022021302024851)

Identification of fertility levels and quantification of the temperature-fertility relationship in rapeseed pol TCMS lines using an improved U-Net++ model

LI Shi-Peng1(), CHEN Cai-Wu2, ZHANG Jing1, LYU Tian1, FU Ting-Dong1, YI Bin1,*()   

  1. 1College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, Hubei, China
    2Guangdong Shaoguan Tobacco Redrying Co., Ltd., Shaoguan 512023, Guangdong, China
  • Received:2024-12-01 Accepted:2025-03-26 Published:2025-06-12 Published online:2025-04-10
  • Contact: *E-mail: yibin@mail.hzau.edu.cn
  • Supported by:
    the National Key Research and Development Program of China(2021YFD1600500);the Wuhan Science and Technology Major Project on Key Techniques of Biological Breeding and Breeding of New Varieties(2022021302024851)

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摘要： 油菜(Brassica napus L.)是全球重要的油料作物, 其杂种优势的利用在提升产量与环境适应能力方面发挥了关键作用。温敏型波里马细胞质雄性不育系(pol TCMS)因其育性受温度影响, 具备一系两用的独特优势, 已成功应用于两系育种。雄蕊、雌蕊长度是生产实践中进行育性等级划分的主要依据, 然而育性等级的主观划分方法易受人为因素干扰, 最终影响表型考察及精细定位的结果。为此, 本研究提出了一种育性等级鉴定的新方法, 即基于改进U-Net++深度学习模型的图像语义分割法。以pol TCMS温敏两系及pol CMS稳定不育系构建F2分离群体, 首先, 获取分离群体中不同育性等级花器官的图像, 标注构建数据集; 其次, 选取U-Net++图像语义分割方法, 通过优化编码器和解码器结构并引入通道注意力模块, 提升模型的分割精度; 最后, 以不同育性等级的图像进行训练和测试。结果表明, 改进后的模型在不同育性等级花器官的分割任务中, 平均交并比为92.02%, 精确率为98.94%, 召回率为98.84%, F1分数为98.87%, 优于其他分割模型方法, 该模型能很好识别出不同育性等级的花器官。基于分割结果获得原位实际长度, 与人工测量的长度相比, 预测值与实测值的决定系数R2为0.989, 均方根误差(RMSE)为0.142 mm, Spearman相关系数为0.983, 可以实现不同育性等级表型参数的准确测量。此外, 通过分析量化温度与育性(雄蕊/雌蕊比值)的关系发现, 育性随单花开花前9 d的温度变化而波动。本研究验证了温度对油菜育性的关键影响, 为深入解析油菜温敏特性及相关基因定位研究提供了新方法和技术支持。

关键词: 油菜, Pol TCMS, U-Net++分割模型, 温度, 育性等级, 基因定位

Abstract:

Rapeseed (Brassica napus L.) is a globally important oil crop, and the exploitation of its hybrid vigor has played a crucial role in enhancing yield and environmental adaptability. The thermosensitive Polima cytoplasmic male sterile (pol TCMS) line has a unique dual-purpose advantage, as its fertility is temperature-dependent, making it highly valuable for two-line breeding systems. In agricultural practice, the classification of fertility status is primarily based on the relative lengths of stamens and pistils. However, traditional fertility grading methods rely on subjective assessment, which is prone to human error, ultimately affecting phenotypic investigations and precise trait mapping. To address this issue, we propose a novel approach for fertility classification based on image semantic segmentation using an improved U-Net++ deep learning model. An F2 segregating population was developed using the pol TCMS thermosensitive line and the pol CMS stable sterile line. First, floral organ images representing different fertility levels within the segregating population were collected and annotated to construct a dataset. Next, we optimized the U-Net++ model by refining its encoder-decoder architecture and integrating a channel attention module to enhance segmentation accuracy. Finally, the model was trained and tested on images corresponding to different fertility levels. Experimental results demonstrated that the improved model achieved a mean intersection-over-union (mIoU) of 92.02%, precision of 98.94%, recall of 98.84%, and an F1 score of 98.87%, outperforming other segmentation models. The model effectively distinguished floral organs across different fertility grades and enabled in situ measurements of organ length based on segmentation results. Compared with manual measurements, the predicted values exhibited a coefficient of determination (R2) of 0.989, a root mean square error (RMSE) of 0.142 mm, and a Spearman correlation coefficient of 0.983, demonstrating high accuracy in phenotypic parameter estimation across different fertility levels. Furthermore, we quantitatively analyzed the relationship between temperature and fertility (stamen/pistil ratio), revealing that fertility fluctuates in response to temperature variations during the nine days preceding anthesis in individual flowers. This study confirms the key role of temperature in regulating fertility and provides a novel methodological framework and technical support for in-depth analyses of temperature-sensitive traits and gene mapping in rapeseed.

Key words: rapeseed, Pol TCMS, U-Net++ segmentation model, temperature, fertility grade, gene location

表1

7级育性分类"

育性等级
Fertility grade		 特征
Trait		 育性分类
Fertility category
0 花药彻底败育, 无花粉或无可育花粉, 花药不能开裂散粉。
The anthers are completely sterile, with no pollen or no fertile pollen, and the anthers cannot dehisce and release pollen.		 0-1级为不育(S)
Levels 0-1 are sterility (S)
1 1-4个雄蕊的花药有微量可育花粉, 但不散粉。
Anthers of 1-4 stamens have trace amounts of fertile pollen but do not shed pollen.
2 5-6个花药有微量可育花粉, 花药膨大部分占正常花药1/3以下, 延迟散粉。
5-6 anthers have trace amounts of fertile pollen, the anther swelling accounts for less than 1/3 of the normal anther, and pollen shedding is delayed.		 2-4级为部分不育(PS)
Levels 2-4 are partial sterility (PS)
3 6个花药有一定数目的可育花粉, 花药膨大部分占正常花药的1/3~1/2。
6 anthers have a certain number of fertile pollen, and the anther swelling accounts for 1/3-1/2 of the normal anther.
4 6个花药有大量可育花粉, 膨大部分占正常花药的1/2-3/4, ♂/♀=4/5。
6 anthers have a large amount of fertile pollen, and the swollen part accounts for 1/2-3/4 of the normal anther, ♂/♀=4/5.
5 6个花药发育正常或基本正常, 花药膨大部分占正常花药的2/3以上, ♂/♀=1。
6 anthers developed normally or almost normally, the anther swelling accounted for more than 2/3 of the normal anther, ♂/♀=1.		 5-6级为可育(F)
Levels 5-6 are fertile (F)
6 6个花药发育正常, ♂/♀>1。
6 anthers developed normally, ♂/♀>1.

图1

油菜花器官图像标注 利用Labelme软件标注花器官图像, 雄蕊标为“stamens”, 雌蕊标为“stigma”。"

图2

改进后的U-Net++网络结构"

表2

2022-2024年温度"

年份(播期)
Years (Sowing date, month/day)		 2月均温
Average temperature
in February (℃)		 3月均温
Average temperature
in March (℃)		 2月负积温
Negative accumulated
temperature in February (℃)		 初花期
Initial flowering
period (month-day)
2021-2022 (10/05) 5.11 15.33 0 03-11
2022-2023 (10/04) 7.75 14.28 0 02-28
2023-2024 (10/08) 5.03 13.63 -8.5 03-04

图3

2022-2024年2月至3月气温"

表3

经典图像分割模型分割效果评价"

模型
Model		 平均交互比
mIoU (%)		 精确率
P (%)		 召回率
R (%)		 F1分数
F1_score (%)
U-Net++ 82.24 97.60 96.95 97.13
U-Kan 86.91 98.39 98.11 98.19
BRAU-Net++ 86.06 98.19 97.84 97.94
Swin-Unet 81.29 97.57 96.90 97.09
ACC-UNet 86.07 98.01 97.92 98.01
STSU-Net 92.02 98.94 98.84 98.87

图4

不同模型的分割效果"

图5

交互界面"

图6

油菜pol TCMS材料育性与雄蕊/雌蕊长度比值相关性 a: 表型参数预测值与实测值关系; b: 花粉数量与雄蕊/雌蕊长度比值关系。"

图7

2022年和2023年盛花期群体育性分布 a~c: 2022年雌蕊、四强雄蕊、雄蕊和雌蕊长度比值频率分布; d~f: 2023年雌蕊、四强雄蕊、雄蕊和雌蕊长度比值频率分布。"

表4

雄蕊/雌蕊比值与温度相关系数"

时段
Time		 相关系数
Correlation coefficient		 时段
Time		 相关系数
Correlation coefficient		 时段
Time		 相关系数
Correlation coefficient		 时段
Time		 相关系数
Correlation coefficient
0- -18 0.63 0- -15 0.73 -3- -12 0.02 0- -6 0.76*
-3- -18 0.18 -3- -15 0.14 -6- -12 -0.13 -3- -6 -0.03
-6- -18 -0.18 -6- -15 -0.04 -9- -12 0.43 0- -3 0.35
-9- -18 -0.07 -9- -15 0.19 0- -9 0.79*
-12- -18 0.01 -12- -15 0.44 -3- -9 -0.34
-15- -18 -0.78 0- -12 0.75 -6- -9 0.59
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