摘要
番木瓜是重要的热带经济水果。番木瓜环斑病毒(Papaya ringspot virus,PRSV)是番木瓜的重要病毒病,经常导致严重的产量损失和质量恶化。自从1998年第一例转基因番木瓜问世以来,使得基于“致病菌衍生的抗病性(pathogen-derived resistance,PDR)”的抗病育种策略获得成功广泛应用。然而依赖于序列同源性的抗病性与病毒突变导致多样性增加之间的矛盾成为番木瓜育种科学家的新挑战。本研究拟采用RNAi策略针对复制酶(nuclear inclusion b.Nib)获得广谱抗PRSV番木瓜新种质。通过团队已建立的胚性愈伤诱导-农杆菌介导转化-再生苗诱导的番木瓜遗传转化体系,共获得经过抗性筛选的再生苗52株,通过特异性PCR进行筛选共计获得24株转基因阳性植株。通过对T0代田间自然发病试验中,转基因番木瓜株系抗病性明显高于非转基因对照,其中NibB5-2田间抗病性最优。通过hiTAIL-PCR方法确定NibB5-2插入位点位于第2号染色体supercontig_30的1976766的位置。T1代接种试验中,无病毒积累且无发病症状,初步确认具有良好的病毒抗性,为番木瓜抗病育种提供新思路。
Papaya is an economically important tropical fruit.Papaya ringspot virus(PRSV)is an important virus disease of papaya,often causing significant yield losses and quality deterioration.Since the introduction of the first transgenic papaya in 1998,PDR-based breeding strategies for disease resistance have been successfully applied.The contradiction between disease resistance based on sequence homology and increased virus genetic diversity became a new challenge for papaya breeding.In this study,we propose to use RNAi strategies aim at nuclear inclusion b gene(Nib)to obtain broad-spectrum resistance to PRSV papaya.With optimized embryo callus generation-Agrobatium meidated transformation-shoot regeneration,52 shoots were obtained after resistance screening and a total of 24 transgenic positive shoots were obtained by specific PCR screening for the T0 generation.In the T0 generation field natural disease test,the transgenic papaya strains were significantly more resistant to disease than the non-transgenic control,with NibB5-2 having the best resistance in the field.Molecular charaterization of the insertion site of Nib5-2 was located at position 1976766 on chromosome 2 supercontig_30 by hiTAIL-PCR method.Green house inoculation test of NibB5-2 T1 generation,there was no virus accumulation and no disease symptoms,confirmed the resistance.Our resulst reveal that RNAi silcencing the Nib gene enhance the papaya virus resistance,meanwhile enriched the candidated genes for virus resistance breeding.
作者
吴清铧
贾瑞宗
郭静远
杨牧之
胡玉娟
郝志刚
赵辉
郭安平
WU Qinghua;JIA Ruizong;GUO Jingyuan;YANG Muzhi;HU Yujuan;HAO Zhigang;ZHAO Hui;GUO Anping(College of Tropical Crops,Hainan University,Haikou,Hainan 570228,China;Hainan Key Laboratory for Biosafety Monitoring and Molecular Breeding in Off-Season Reproduction Regions/Sanya Research Institutey,Chinese Academy of Tropical Agricultural Sciences/Institute of Tropical Bioscience and Biotechnology,Chinese Academy of Tropical Agricultural Sciences,Sanya,Hainan 572024,China)
出处
《热带作物学报》
CSCD
北大核心
2024年第4期837-846,共10页
Chinese Journal of Tropical Crops
基金
海南省重大科技计划项目(No.ZDKJ202002)
海南省重点研发计划项目(No.ZDYF2022XDNY257)
崖州湾科技城菁英人才项目(No.SCKJ-JYRC-2022-67)。
