果蔗中甘蔗线条花叶病毒脱除技术

doi:10.13560/j.cnki.biotech.bull.1985.2025-0268

生物技术通报 ›› 2025, Vol. 41 ›› Issue (9): 62-70.doi: 10.13560/j.cnki.biotech.bull.1985.2025-0268

果蔗中甘蔗线条花叶病毒脱除技术

胡鑫¹^,²(), 罗正英³, 刘新龙¹^,², 吴才文¹^,², 吴转娣¹^,²()

^1.云南省农业科学院热带作物生物育种全国重点实验室，昆明 650205
^2.云南省农业科学院甘蔗研究所云南甘蔗遗传改良重点实验室农业农村部甘蔗生物学与遗传育种重点实验室（云南），开远 661699
^3.云南农业大学农学与生物技术学院云南省作物生产与智慧农业重点实验室，昆明 650201

收稿日期:2025-03-13 出版日期:2025-09-26 发布日期:2025-09-24
通讯作者: 吴转娣，女，硕士，副研究员，研究方向：甘蔗健康种苗与生物技术；E-mail: wzd@yaas.org.cn
作者简介:胡鑫，男，博士，助理研究员，研究方向：甘蔗生物育种；E-mail: sugarhuxin@163.com
基金资助:
国家自然科学基金项目(32460740);云南省农业科学院科研项目(2024KYZ X-03);云南省种子种业联合实验室项目(202205AR070001-09)

Elimination Technology of Sugarcane Streak Mosaic Virus in Chewing Cane

HU Xin¹^,²(), LUO Zheng-ying³, LIU Xin-long¹^,², WU Cai-wen¹^,², WU Zhuan-di¹^,²()

^1.National Key Laboratory for Biological Breeding of Tropical Crops, Yunnan Academy of Agricultural Sciences, Kunming 650205
^2.Sugarcane Research Institute, Yunnan Academy of Agricultural Sciences, Yunnan Key Laboratory of Sugarcane Genetic Improvement, Key Laboratory of Sugarcane Biology and Genetic Breeding (Yunnan), Ministry of Agriculture and Rural Affairs, Kaiyuan 661699
^3.College of Agronomy and Biotechnology, Yunnan Agricultural University, The Key Laboratory for Crop Production and Smart Agriculture of Yunnan Province, Kunming 650201

Received:2025-03-13 Published:2025-09-26 Online:2025-09-24

摘要/Abstract

摘要：

目的研究果蔗‘Badila’中甘蔗线条花叶病毒（sugarcane streak mosaic virus, SCSMV）的高效脱毒技术，为提升果蔗产量提供技术支撑。方法以携带SCSMV的‘Badila’为试验材料，设置未处理对照、50 ℃ 2 h、50 ℃ 3 h、52 ℃ 3 h、54 ℃ 3 h和58 ℃ 1 h共6个温水处理组，结合茎尖组织培养和芽尖组织培养生产脱毒苗，并对脱毒苗进行SCSMV病原分子检测，同时调查脱毒苗桶栽后的农艺性状，综合分析不同处理对SCSMV的脱除效果。结果温水处理显著降低蔗芽出芽率，且温度和处理时间与出芽率呈负相关。当温度升至54 ℃和58 ℃时，蔗芽完全丧失萌发能力，导致无法获取茎尖组织培养材料；然而，该处理对芽分生组织培养的成活率影响较小，即使在54 ℃和58 ℃条件下，芽尖分生组织仍保持较高的成活率（83.3%和76.9%）。在病毒脱除效果方面，50 ℃ 2 h、50 ℃ 3 h、52 ℃ 3 h温水处理结合茎尖分生组织培养仅能降低SCSMV检出率至80%，而相同条件下芽尖培养则分别降至70%、60%和30%。特别是在58 ℃处理1 h条件下，芽尖培养的病毒脱除率可达100%。田间试验表明，脱毒处理的甘蔗植株在长势和农艺性状方面均显著优于对照，其中，株高、中部茎节间长度、单茎重、锤度和蔗糖分分别提高43.5%、53.5%、58.3%、12.4%和30.0%。结论成功建立了温水处理结合芽尖组织培养的SCSMV脱除技术体系，该技术具有成活率高、脱毒效率高等优点。

关键词: 果蔗, SCSMV, 热处理, 芽尖组织培养, 脱毒

Abstract:

Objective This study is aimed to investigate efficient virus elimination technology for Sugarcane Streak Mosaic Virus (SCSMV) in chewing cane ‘Badila’, providing technical support for improving sugarcane yield. Method Using SCSMV-infected ‘Badila’ as experimental material, six hot water treatment groups were established: Untreated control, 50 ℃ for 2 h, 50 ℃ for 3 h, 52 ℃ for 3 h, 54 ℃ for 3 h, and 58 ℃ for 1 h. Virus-free plantlets were produced through shoot-tip culture and bud-tip culture. The virus-free plantlets were subjected to molecular detection of SCSMV, and their agronomic traits were investigated after pot cultivation to comprehensively analyze the virus elimination effects. Result Hot water treatment significantly reduced bud germination rates, with temperature and treatment duration showing a negative correlation with germination rate. No sprouting was observed at 54 ℃ and 58 ℃, making it impossible to obtain shoot-tip explants for tissue culture. However, the treatment had minimal impact on the survival rate of bud-tip culture. Even at 54 ℃ and 58 ℃, the bud apical meristems still maintained relatively high survival rates (83.3% and 76.9%, respectively). In terms of virus elimination, hot water treatment at 50 ℃ for 2 h, 50 ℃ for 3 h, and 52 ℃ for 3 h combined with shoot-tip culture only reduced the SCSMV detection rate to 80%. Under the same conditions, bud-tip culture reduced the rates to 70%, 60%, and 30%, respectively. Notably, under 58 ℃ treatment for 1 h, bud-tip culture achieved complete virus elimination (100%). Field trials demonstrated that virus-free sugarcane plants exhibited significantly better growth and agronomic traits compared to the control group. Plant height, internode length, single stem weight, brix and sugar content increased by 43.5%, 53.5%, 58.3%, 12.4% and 30.0%, respectively. Conclusion This study successfully established a virus elimination system combining hot water treatment and bud-tip culture, characterized by high plantlet regeneration and virus elimination efficiency.

Key words: chewing cane, SCSMV, heat treatment, bud-tip culture, virus elimination

胡鑫, 罗正英, 刘新龙, 吴才文, 吴转娣. 果蔗中甘蔗线条花叶病毒脱除技术[J]. 生物技术通报, 2025, 41(9): 62-70.

HU Xin, LUO Zheng-ying, LIU Xin-long, WU Cai-wen, WU Zhuan-di. Elimination Technology of Sugarcane Streak Mosaic Virus in Chewing Cane[J]. Biotechnology Bulletin, 2025, 41(9): 62-70.

图/表 7

图1 果蔗芽尖分生组织培养技术流程A：温水处理；B：剥除芽鳞；C：取芽尖组织；D：芽尖组织接种到MS培养基中；E：芽尖组织分化成幼苗；F：幼苗继代培养成丛生苗

Fig. 1 Flowchart of the bud-tips culture technique of chewing caneA: Hot water treatment. B: Removing bud scales. C: Extracting bud tip. D: Inoculation of bud-tip onto MS medium. E: Differentiation of bud-tip into seedlings. F: Subculture of seedlings into clumping seedlings

图2 果蔗RNA的提取和SCSMV分子检测A：提取果蔗叶片总RNA；B：PCR检测SCSMV病毒；M为DNA分子marker，N为以无菌水阴性对照；P为携带SCSMV病毒的cDNA阳性对照，1‒20为样品编号

Fig. 2 RNA extraction and molecular detection of SCSMV in chewing caneA: Extraction of total RNA from chewing cane leaves. B: PCR detection of SCSMV virus. M indicates the DNA molecular marker, N is the negative control using sterile water, P is the positive control carrying SCSMV virus, and samples are numbered 1‒20

图3 PCR产物序列同源性分析

Fig. 3 Homology analysis of PCR product sequences

表1 不同温水处理条件对茎芽存活率的影响

Table 1 Effects of different hot water treatment conditions on the survival rates of stem buds

编号 No.	温水处理条件 Hot water treatment conditions					芽尖组织培养 Bud-tip culture
编号 No.	温水处理条件 Hot water treatment conditions	出芽率Germination rate (%)	茎尖成活率Survival rate of stem tips (%)	污染率 Contamination rate (%)	芽苗状态 Emergence of seedlings	芽尖成活率 Survival rate of bud-tips (%)	污染率 Contamination rate (%)	芽苗状态 Emergence of seedlings
A	未处理	86.3	81.8	15.7	丛生芽，长势良好	87.0	11.9	丛生芽，长势良好
B	50 ℃ 2 h	37.5	89.3	10.0	丛生芽，长势良好	95.2	5.0	丛生芽，长势良好
C	50 ℃ 3 h	23.8	85.7	13.2	丛生芽，长势良好	90.9	5.6	丛生芽，长势良好
D	52 ℃ 3 h	12.9	84.6	15.0	丛生芽，长势良好	90.9	3.1	丛生芽，长势良好
E	54 ℃ 3 h	0	-		-	83.3	2.3	丛生芽，长势良好
F	58 ℃ 1 h	0	-		-	76.9	0	丛生芽，长势良好

图4 不同处理条件下果蔗组培苗的SCSMV脱毒分子检测A：未温水处理组；B：50 ℃ 2 h处理组；C：50 ℃ 3 h处理组；D：52 ℃ 3 h处理组；E：54 ℃ 3 h处理组；F：58 ℃ 1 h处理组。A‒D中，1‒10为茎尖组培苗样品，11‒20为芽尖组培苗样品。E‒F中，1‒20为芽尖组培样品。M：DNA分子marker；N：无菌水阴性对照；P：携带SCSMV病毒的cDNA阳性对照

Fig. 4 SCSMV molecular detection in tissue-cultured seedlings of chewing cane under different treatment conditionsA: Non-hot water treatment group. B: The 50 ℃ 2 h hot water treatment group. C: The 50 ℃ 3 h hot water treatment group. D: The 52 ℃ 3 h hot water treatment group. E: The 54 ℃ 3 h treatment group. F: The 58 ℃ 1 h treatment group. A‒D: Samples 1‒10 are from stem-tip culture seedlings, and samples 11‒20 are from bud-tip culture seedlings. E‒F: Sample 1‒20 were from bud tip culture seedlings. M: DNA molecular marker. N: Sterile water negative control. P: Positive control with SCSMV virus cDNA

图5 脱毒果蔗和非脱毒果蔗的线条花叶病症状及农艺性状

Fig. 5 Symptoms of mosaic disease and agronomic traits of virus-infected and virus-free chewing cane

表2 非脱毒果蔗与脱毒果蔗的农艺性状比较

Table 2 Comparison of agronomic traits between virus-infected and virus-free chewing cane

材料

Material

株高

Plant height (cm)

茎径

Stem diameter (cm)

节间长度

Internode length (cm)

有效茎

Effective stems (Stems/pot)

ingle stem weight (kg)

单茎重 S

锤度

Brix (%)

蔗糖分

Sucrose content (%)

非脱毒果蔗

Virus-infected chewing cane

脱毒果蔗

Virus-free chewing cane

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果蔗中甘蔗线条花叶病毒脱除技术

Elimination Technology of Sugarcane Streak Mosaic Virus in Chewing Cane

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