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作物学报 ›› 2025, Vol. 51 ›› Issue (11): 2983-2995.doi: 10.3724/SP.J.1006.2025.55029

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蓝型油菜磷转运蛋白BnaPT48的功能研究

张雯(), 李玉, 王创, 石磊, 丁广大()   

  1. 华中农业大学资源与环境学院 / 微量元素研究中心 / 农业农村部长江中下游耕地保育重点实验室, 湖北武汉 430070
  • 收稿日期:2025-04-18 接受日期:2025-08-13 出版日期:2025-11-12 网络出版日期:2025-08-19
  • 通讯作者: *丁广大, E-mail: dgd@mail.hzau.edu.cn
  • 作者简介:E-mail: zhangw@webmail.hzau.edu.cn
  • 基金资助:
    农业生物育种国家科技重大专项(2023ZD04072)

Functional study of the phosphate transporter protein BnaPT48 in Brassica napus

ZHANG Wen(), LI Yu, WANG Chuang, SHI Lei, DING Guang-Da()   

  1. College of Resources and Environment, Huazhong Agricultural University / Microelement Research Center / Key Laboratory of Arable Land Conservation in the Middle and Lower Reaches of Yangtze River, Ministry of Agriculture and Rural Affairs, Wuhan 430070, Hubei, China
  • Received:2025-04-18 Accepted:2025-08-13 Published:2025-11-12 Published online:2025-08-19
  • Contact: *E-mail: dgd@mail.hzau.edu.cn
  • Supported by:
    National Key R&D Program for Agricultural Biological Breeding(2023ZD04072)

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摘要：

磷是植物生长发育必需的大量营养元素之一。油菜作为我国重要的油料作物, 具有需磷多且对缺磷敏感的特性。本研究以在甘蓝型油菜根系中表达丰度高且受缺磷诱导的基因BnaPT48为研究对象, 通过分析其基因表达模式、蛋白亚细胞定位及遗传材料的表型等, 揭示该基因在油菜磷吸收中的作用。研究发现BnaPT48定位于细胞膜, 且在根系各组织中受缺磷显著诱导表达。BnaPT48能回补拟南芥磷吸收突变体(atpt1/2)的表型, 表明BnaPT48能促进植物的磷吸收。在拟南芥中过表达BnaPT48能促进植株在正常磷和低磷条件下的生长。在油菜中过表达BnaPT48, 低磷处理后显著增加地上部干重, 降低子叶中的无机磷浓度, 促进子叶中叶绿素的降解和子叶衰老, 根系无机磷浓度在正常磷时显著增加。本研究揭示了BnaPT48在调控植物磷吸收中的作用, 为甘蓝型油菜磷效率的遗传改良提供了理论基础和基因资源。

关键词: 甘蓝型油菜, BnaPT48, 转运蛋白, 低磷胁迫, 磷浓度, 磷吸收

Abstract:

Phosphorus (P) is an essential macronutrient required for plant growth and development. Rapeseed (Brassica napus), a major oilseed crop in China, is characterized by a high demand for P and strong sensitivity to P deficiency. This study investigates the gene BnaPT48, which is highly expressed in the root system of Brassica napus and is strongly induced under P-deficient conditions. Through analyses of its expression pattern, protein subcellular localization, and phenotypes of transgenic materials, the role of BnaPT48 in P uptake was elucidated. BnaPT48 was found to localize to the plasma membrane and was significantly upregulated by P deficiency in various root tissues. The BnaPT48 protein complemented the P uptake defect of the Arabidopsis thaliana mutant atpt1/2, indicating its functional role in enhancing P acquisition. Overexpression of BnaPT48 in A. thaliana promoted plant growth under both normal and low P conditions. In rapeseed, overexpression of BnaPT48 significantly increased the shoot dry weight under low P treatment, reduced inorganic P concentration in cotyledons, accelerated chlorophyll degradation and cotyledon senescence, and increased inorganic P accumulation in roots under normal P conditions. These findings reveal the function of BnaPT48 in regulating phosphate uptake and redistribution, and provide a theoretical foundation and genetic resource for improving P use efficiency in B. napus through genetic approaches.

Key words: Brassica napus, BnaPT48, transport protein, low phosphorus stress, P concentration, P absorption

表1

本研究中使用的引物序列"

引物名称
Primer name		 引物序列
Primer sequence (5′-3′)		 引物作用
Function of primers
BnaPT48-GFP-F ATCGATTCTAGAGCC ATGGCTGATAATCAGCAGCTAGGAG 构建亚细胞定位载体
Construct a subcellular localization vector
BnaPT48-GFP-R TTCTCCTTTGCCCAT GTTTCTTCCGGAGGTAG
BnaPT48p-GUS-F CAGGTCGACGGATCCATATCAACTTTATGGAAAGATGACAA 构建组织定位载体
Construct a tissue-specific targeting vector
BnaPT48p-GUS-R TAAGGGACTGACCACTTGCCTTCTTCTAGATTATCTACA
BnPT48OX-F CCCCCGGG ATGGCTGATAATCAGCAGCTAGGAG 构建过表达载体
Construct an overexpression vector
BnPT48OX-R GCGTCGAC TCAGTTTCTTCCGGAGGTAG
BnaPT48-qPCR-F TATTATTGGCGAATGAAGATGC 定量基因表达水平
Quantify gene expression levels
BnaPT48-qPCR-R CCTTGGAGAACAAGCCGTAG

图1

BnaPT48及其同源基因的进化及序列相似性分析 A: 系统发育关系分析; B: 氨基酸序列比对; C: 蛋白三维结构分析。Merge: AtPHT1;3和BnaPT48融合。"

图2

BnaPT48的时空表达模式 A: 正常磷和低磷处理下苗期根中BnaPT48表达分析(以正常磷处理0 h根部的表达量为对照); B: 正常磷(CK)和低磷(LP)处理下苗期地上部BnaPT48表达分析(以正常磷处理0 h地上部的表达量为对照); C: 苗期不同组织中BnaPT48表达模式分析(以正常磷处理0 h苗期不同组织的表达量为对照); D: 成熟期不同组织中BnaPT48表达模式分析(以正常磷处理0 h成熟期不同组织的表达量为对照; CK: 90 kg hm-2; LP: 15 kg hm-2)。FEL: 完全展开叶; CL: 茎生叶; IS: 花序茎; B: 花蕾; F: 花; Hu: 角果壳; S: 种子; Ca: 角果柄。苗期, CK: 1 mmol L-1 P; -P: 0 mmol L-1 P。数据表示为平均值±标准差(n = 3)。t检验。*、**、***分别表示在0.05、0.01和0.001水平差异显著。"

图3

BnaPT48蛋白的亚细胞定位和组织定位 A: BnaPT48蛋白的亚细胞定位; B: BnaPT48p::GUS的转基因材料GUS染色, a~g依次为正常磷(CK: 1 mmol L-1 P)条件下的根系(a), 低磷(LP: 5 μmol L-1 P)条件下的根系(b)、子叶(c)、真叶(d)、下胚轴(e)、花(f)和角果(g), a~b标尺为0.5 mm, c~g标尺为0.2 cm。"

图4

BnaPT48互补拟南芥磷吸收突变体atpt1/2材料在不同磷水平下的表型 A: 拟南芥回补材料的鉴定; B: 拟南芥过表达BnaPT48材料和野生型Col-0的长势; C: 正常磷和低磷条件下的鲜重; D: 正常磷和低磷条件下的无机磷浓度。t检验(Student’s t-test)。*、**、***分别表示在0.05、0.01和0.001水平差异显著。CK: 正常磷处理(625 μmol L-1 P); LP: 低磷处理(15 μmol L-1 P)。B图标尺为0.5 cm。以相同条件下的野生型Col-0为对照。"

图5

拟南芥过表达BnaPT48和野生型材料在不同磷水平下的表型 A: 拟南芥过表达BnaPT48材料的鉴定; B: 拟南芥过表达BnaPT48材料和野生型Col-0的长势; C: 根系和地上部鲜重; D: 根系和地上部干重; E: 根系和地上部无机磷浓度; F: 地上部和根系总磷浓度; G: 总根长; H: 根系总表面积; I: 地上部和根系磷累积量。数据表示为平均值±标准差(n = 3)。t检验(Student’s t-test)。*、**、***分别表示在0.05、0.01和0.001水平差异显著。CK: 正常磷处理(200 μmol L-1 P); LP: 低磷处理(5 μmol L-1 P)。B图标尺为3 cm。以相同条件下的野生型Col-0为对照。"

图6

油菜过表达BnaPT48和野生型材料在不同磷水平下的表型 A: 各材料在正常磷条件下的长势; B: 各材料在低磷条件下的长势; C: 油菜过表达BnaPT48材料的鉴定; D: 正常磷和低磷条件下的干重; E: 正常磷和低磷条件下的无机磷浓度; F: 地上部磷分配比例; G: 正常磷和低磷条件下的总磷浓度; H: 磷转运系数; I: 正常磷和低磷条件下的磷累积量。标尺= 5 cm。a代表子叶(cotyledon), b代表第1片真叶(leaf1)。数据表示为平均值±标准差(n = 3)。t检验, *、**、***分别表示在0.05、0.01和0.001水平差异显著。CK: 正常磷处理(1 mmol L-1 P); LP: 低磷处理(10 μmol L-1 P)。以相同条件下的野生型Col-0为对照。"

图7

过表达BnaPT48对油菜叶片叶绿素和类胡萝卜素含量的影响 A: 正常磷和低磷条件下的叶绿素a含量; B: 正常磷和低磷条件下的叶绿素b含量; C: 正常磷和低磷条件下的类胡萝卜素含量; D: 正常磷和低磷条件下的总叶绿素含量; E: 正常磷和低磷条件下的SPAD值。数据表示为平均值±标准差(n = 3), t检验。*、**、***分别表示在0.05、0.01和0.001水平差异显著。CK: 正常磷处理(1 mmol L-1 P); LP: 低磷处理(10 μmol L-1 P)。以相同条件下的野生型Col-0为对照。"

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