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A Strategy for the Production and Molecular Validation of Agrobacterium-Mediated Intragenic Octoploid Strawberry

文献类型： 外文期刊

作者： Duan, Ke 1 ; Zhao, Ying-Jie 2 ; Li, Zi-Yi 3 ; Zou, Xiao-Hua 1 ; Yang, Jing 1 ; Guo, Cheng-Lin 4 ; Chen, Si-Yu 5 ; Yang, Xiu-Rong 1 ; Gao, Qing-Hua 1 ;

作者机构： 1.Shanghai Acad Agr Sci SAAS, Shanghai Key Lab Protected Hort Technol, Forestry & Fruit Tree Res Inst, Shanghai 201403, Peoples R China

2.Lanzhou New Area Acad Modern Agr Sci, Lanzhou 730300, Peoples R China

3.Shanghai Inst Technol, Ecol Tech & Engn Coll, Shanghai 201418, Peoples R China

4.Hangzhou Woosen Biotechnol Co Ltd, Hangzhou 310012, Peoples R China

5.Shanghai Ocean Univ, Coll Food Sci, Shanghai 201306, Peoples R China

关键词： Fragaria x ananassa; sugar transporter; intragenesis; Agrobacterium-mediated transformation; droplet digital PCR

期刊名称：PLANTS-BASEL （ 影响因子：4.658； 五年影响因子：4.827 ）

ISSN：

年卷期： 2021 年 10 卷 11 期

页码：

收录情况： SCI

摘要： Intragenesis is an all-native engineering technology for crop improvement. Using an intragenic strategy to bring genes from wild species to cultivated strawberry could expand the genetic variability. A robust regeneration protocol was developed for the strawberry cv. 'Shanghai Angel' by optimizing the dose of Thidiazuron and identifying the most suitable explants. The expression cassette was assembled with all DNA fragments from F. vesca, harboring a sugar transporter gene FvSTP8 driven by a fruit-specific FvKnox promoter. Transformed strawberry was developed through an Agrobacterium-mediated strategy without any selectable markers. Other than PCR selection, probe-based duplex droplet digital PCR (ddPCR) was performed to determine the T-DNA insert. Four independent transformed shoots were obtained with a maximum of 5.3% efficiency. Two lines were confirmed to be chimeras, while the other two were complete transformants with six and 11 copies of the intragene, respectively. The presence of a vector backbone beyond the T-DNA in these transformants indicated that intragenic strawberries were not obtained. The current work optimized the procedures for producing transformed strawberry without antibiotic selection, and accurately determined the insertion copies by ddPCR in the strawberry genome for the first time. These strategies might be promising for the engineering of 'Shanghai Angel' and other cultivars to improve agronomic traits.

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