中国农业科学 ›› 2022, Vol. 55 ›› Issue (19): 3767-3778.doi: 10.3864/j.issn.0578-1752.2022.19.007
肖桂华1,2,3(),文康1,2,3,韩健4,郝晨星1,2,3,叶蓉春1,2,3,朱亦赤1,2,3,萧顺元1,邓子牛1,2,3,马先锋1,2,3(
)
收稿日期:
2022-04-11
接受日期:
2022-05-17
出版日期:
2022-10-01
发布日期:
2022-10-10
通讯作者:
马先锋
作者简介:
肖桂华,E-mail: 基金资助:
XIAO GuiHua1,2,3(),WEN Kang1,2,3,HAN Jian4,HAO ChenXing1,2,3,YE RongChun1,2,3,ZHU YiChi1,2,3,XIAO ShunYuan1,DENG ZiNiu1,2,3,MA XianFeng1,2,3(
)
Received:
2022-04-11
Accepted:
2022-05-17
Online:
2022-10-01
Published:
2022-10-10
Contact:
XianFeng MA
摘要:
【背景】柑橘溃疡病是由柑橘黄单胞杆菌柑橘致病变种(Xanthomonas citri subsp. citri,Xcc)引起的细菌性病害,可侵染枝、叶、果,危害几乎所有的柑橘主栽品种。前期对湖南省39个柑橘园的调查结果显示柑橘果园土壤酸化和交换性钙缺乏严重,叶片中均存在钙缺乏现象。钙是植物所需大量元素之一,钙缺乏会造成植物营养失衡,生长势下降,植物免疫水平受影响。然而,钙元素对柑橘溃疡病抗性的影响尚不明确。【目的】分析对柑橘溃疡病敏感的枳(Poncirus trifoliata)在不同钙浓度处理下叶片注射接种Xcc后的致病差异,探讨钙在Xcc侵染枳叶片过程中的作用。【方法】采用沙培法对枳实生苗进行0、0.75、3、30 mmol·L-1钙浓度处理,分别测定枳生长期的生物量、叶绿素a和b浓度、根系和叶片钙元素含量、观察根系活性氧(ROS)的产生和胼胝质沉积,并分析枳叶片接种Xcc后细胞壁合成相关基因及免疫途径相关基因诱导表达变化特征。【结果】以3 mmol·L-1钙处理为对照,0、0.75和30 mmol·L-1钙处理后,枳地上部和地下部生长发育均受抑制,叶绿素a和b浓度降低;根系与叶片中的钙含量与外源钙施加量成正比;不同钙浓度处理后根系中产生ROS和胼胝质沉积,在3 mmol·L-1处理时达到最大值;枳叶片接种Xcc后,随着钙浓度增加,叶片症状逐渐减轻,但Xcc的生长量无明显差异;相较于3 mmol·L-1处理,参与细胞壁合成相关基因PtCESA4在0 mmol·L-1处理下受Xcc诱导先上调表达后下调表达,在30 mmol·L-1处理下受Xcc诱导上调表达,PtPME和PtFLA在0 mmol·L-1处理下受Xcc诱导下调表达,在30 mmol·L-1处理下受Xcc诱导上调表达;叶片接种Xcc 0、2、4、6 dpi后免疫途径相关基因PtGSL、PtGST1、PtWRKY22在30 mmol·L-1处理下受Xcc诱导表达水平高于0 和3 mmol·L-1处理。【结论】钙缺乏和过量均会影响枳生长发育,引起叶片失绿,根系产生ROS和胼胝质沉积均有所减少。施钙后接种Xcc,叶片表面的感病症状明显减弱,但菌含量与对照无显著差异。钙可能通过调控细胞壁合成相关基因促使细胞壁增厚,从而抑制Xcc突破叶片表皮形成典型症状。
肖桂华,文康,韩健,郝晨星,叶蓉春,朱亦赤,萧顺元,邓子牛,马先锋. 钙对枳生长发育及柑橘溃疡病抗性的影响[J]. 中国农业科学, 2022, 55(19): 3767-3778.
XIAO GuiHua,WEN Kang,HAN Jian,HAO ChenXing,YE RongChun,ZHU YiChi,XIAO ShunYuan,DENG ZiNiu,MA XianFeng. Effects of Calcium on Growth and Development of Poncirus trifoliata and Resistance to Citrus Canker[J]. Scientia Agricultura Sinica, 2022, 55(19): 3767-3778.
表1
PTI免疫响应相关基因及细胞壁合成相关基因的引物列表"
基因Gene | 引物名称Primer name | 引物序列Primer sequence (5′-3′) | Tm (℃) |
---|---|---|---|
PtGSL | PtGSL-qPCR-F | GGTGGGATGGAGAACAAGAACACC | 59.8 |
PtGSL-qPCR-R | GGTACCAAATCTTCGTCTGCCCAT | 59.3 | |
PtWAKY22 | PtWAKY22-qPCR-F | GCGGATTGTCTCGCATGTG | 59.6 |
PtWAKY22-qPCR-R | TTATGGGTTTCTGCCCGTATTT | 60.0 | |
PtGST1 | PtGST1-qPCR-F | GCCCGTTTGTCTCAGTCCAA | 59.8 |
PtGST1-qPCR-R | TGCAAATCGACCAAGGTGAA | 59.7 | |
PtCESA4 | PtCESA4-qPCR-F | GATGGGCTCTTGGCTCTGTT | 59.2 |
PtCESA4-qPCR-R | GGTGTTGGTGTATGCAAGCC | 59.4 | |
Actin | Actin-qPCR-F | CACACTGGAGTGATGGTTGG | 59.4 |
Actin-qPCR-R | ATTGGCCTTGGGGTTAAGAG | 60.0 | |
PtPME | PtPME-qPCR-F | GAACCTAACGGAAGCCCACA | 58.6 |
PtPME-qPCR-R | GACTCCGTTGCTCGACTTCA | 59.3 | |
PtFLA | PtFLA-qPCR-F | GACCCTCTACCCGAACCTCT | 59.4 |
PtFLA-qPCR-R | GCAGCGTCATGTTGAACGAA | 59.1 | |
PtRbohD | PtRbohD-qPCR-F | GTAAATTGCGCTGCCGTCTC | 58.3 |
PtRbohD-qPCR-R | GTCCAATCGCCCAAAGTTCG | 59.2 | |
PtRbohF | PtRbohF-qPCR-F | GACCTTGTCAAAGGGGCAGA | 59.6 |
PtRbohF-qPCR-R | CCTATCGAAGGGCTTTGGCA | 58.1 |
图7
不同钙浓度处理下枳根系DAB和胼胝质染色 A:枳根系DAB染色,比例尺:200 μm DAB staining of P. trifoliata roots, scale: 200 μm;B:枳根系的胼胝质染色,比例尺:100 μm Callose staining of P. trifoliata roots, scale: 100 μm;C、D:PtRbohD、PtRbohF相对表达量Relative expression level of PtRbohD, PtRbohF;E:胼胝质沉积部位平均荧光强度Average fluorescence intensity of callose deposition site"
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