Identification of two plastid transit peptides for construction of pollen-inactivation system in rice

IF 2.6 3区 农林科学 Q1 AGRONOMY
Menglong Wang, Xiaoqun Peng, Changjian Wang, Xiaoyan Tang
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Abstract

Hybrid seed production technology (SPT) is achieved through the utilization of a recessive nuclear male-sterile mutant transformed with a transgenic cassette comprising three essential components: the wild-type gene to restore the fertility of the male-sterile mutant, an α-amylase gene to disrupt transgenic pollen grains, and red fluorescence protein gene DsRed to distinguish the transgenic seeds from the nontransgenic male sterile seeds. In rice, we establish the pollen disruption system by introducing an amyloplast targeting signal peptide (ASP) at the N-terminus of maize α-amylase protein ZM-AA1ΔSP (ZM-AA1 with the N-terminal signal peptide removed). The ASP facilitates the transport of ZM-AA1ΔSP protein into amyloplast where it degrades starch, resulting in disruption of the pollen fertility. To obtain such signal peptides for rice, we searched the rice proteins homologous to the defined wheat amyloplast proteins followed by protein–protein interaction network predictions and targeting signal peptides prediction. These analyses enabled the identification of four candidate ASPs in rice, which were designated as ASP1, ASP2, ASP3, and ASP4, respectively. ASP1 and ASP2, when linked with ZM-AA1ΔSP, exhibited the capability to disrupt transgenic pollen grains, whereas ASP3 and ASP4 did not produce this effect. Interestingly, the localization experiments showed that ASP3 and ASP4 were able to target the proteins into chloroplast. The ASP1 and ASP2 sequences provide valuable tools for genetic engineering of the rice male-sterile system, which will contribute to the hybrid rice breeding and production.

Abstract Image

鉴定用于构建水稻花粉失活系统的两种质粒转运肽
杂交种子生产技术(SPT)是通过利用隐性核雄性不育突变体转化转基因盒来实现的,转基因盒由三个重要部分组成:野生型基因用于恢复雄性不育突变体的育性;α-淀粉酶基因用于破坏转基因花粉粒;红色荧光蛋白基因DsRed用于区分转基因种子和非转基因雄性不育种子。在水稻中,我们通过在玉米α-淀粉酶蛋白ZM-AA1ΔSP(去除N端信号肽的ZM-AA1)的N端引入一个淀粉质靶向信号肽(ASP)来建立花粉干扰系统。α-淀粉酶蛋白 ZM-AA1ΔSP (ZM-AA1,去除了 N 端信号肽)有助于 ZM-AA1ΔSP 蛋白转运到淀粉质中,在淀粉质中降解淀粉,从而破坏花粉的繁殖力。为了获得水稻的此类信号肽,我们搜索了与已定义的小麦淀粉体蛋白同源的水稻蛋白,然后进行了蛋白-蛋白相互作用网络预测和靶向信号肽预测。通过这些分析,确定了水稻中的四个候选 ASP,分别命名为 ASP1、ASP2、ASP3 和 ASP4。ASP1和ASP2与ZM-AA1ΔSP连接后能破坏转基因花粉粒,而ASP3和ASP4则不会产生这种效果。有趣的是,定位实验表明,ASP3 和 ASP4 能够将蛋白质定位到叶绿体中。ASP1 和 ASP2 序列为水稻雄性不育系统的基因工程提供了宝贵的工具,将有助于杂交水稻的育种和生产。
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来源期刊
Molecular Breeding
Molecular Breeding 农林科学-农艺学
CiteScore
5.60
自引率
6.50%
发文量
67
审稿时长
1.5 months
期刊介绍: Molecular Breeding is an international journal publishing papers on applications of plant molecular biology, i.e., research most likely leading to practical applications. The practical applications might relate to the Developing as well as the industrialised World and have demonstrable benefits for the seed industry, farmers, processing industry, the environment and the consumer. All papers published should contribute to the understanding and progress of modern plant breeding, encompassing the scientific disciplines of molecular biology, biochemistry, genetics, physiology, pathology, plant breeding, and ecology among others. Molecular Breeding welcomes the following categories of papers: full papers, short communications, papers describing novel methods and review papers. All submission will be subject to peer review ensuring the highest possible scientific quality standards. Molecular Breeding core areas: Molecular Breeding will consider manuscripts describing contemporary methods of molecular genetics and genomic analysis, structural and functional genomics in crops, proteomics and metabolic profiling, abiotic stress and field evaluation of transgenic crops containing particular traits. Manuscripts on marker assisted breeding are also of major interest, in particular novel approaches and new results of marker assisted breeding, QTL cloning, integration of conventional and marker assisted breeding, and QTL studies in crop plants.
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