Cloning and Expression Analysis of Different Truncated U3 Promoters in Phyllostachys edulis
Huijin Fan1,2, Kangming Jin1, Renying Zhuo1, Guirong Qiao,1,*1The Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China 2Nanjing Forestry University, Nanjing 210037, China
Abstract The U3 and U6 promoters with well-defined transcription initiation sites are important elements driving sgRNA transcription in the CRISPR/Cas9 genome editing system. According to the two sequences of PeU3 promoter cloned from Phyllostachys edulis, six different truncated U3 promoters were successfully cloned and were 550 bp, 397 bp, 149 bp, and 561 bp, 392 bp, 152 bp, respectively in length. GUS and LUC expression vectors were constructed by corresponding truncated promoter and transformed into the callus of Dendrocalamus latiflorus and tobacco leaf by the Agrobacterium-mediated method, respectively. Our results indicate that, all of these U3 promoters have different transcriptional activity, and the Peu3-1-2pro promoter with a length of 397 bp has the strongest activity. It provides more ideal endogenous promoters for constructing CRISPR/Cas9 genome editing system of P. edulis. Keywords：genome editing;LUC activity;PeU3 promoter;Phyllostachys edulis
PDF (19220KB)摘要页面多维度评价相关文章导出EndNote|Ris|Bibtex收藏本文 引用本文 凡惠金, 金康鸣, 卓仁英, 乔桂荣. 毛竹不同截短U3启动子的克隆及表达分析. 植物学报, 2020, 55(3): 299-307 doi:10.11983/CBB19217 Fan Huijin, Jin Kangming, Zhuo Renying, Qiao Guirong. Cloning and Expression Analysis of Different Truncated U3 Promoters in Phyllostachys edulis. Chinese Bulletin of Botany, 2020, 55(3): 299-307 doi:10.11983/CBB19217
竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点。毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018)。近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用。基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013)。基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的
利用毛竹基因组在NCBI网站进行blast, 获得多个U3 snRNA序列, 并对其进行启动子元件分析, 然后设计并合成引物(表1)。以毛竹叶片基因组DNA为模板, 扩下划线部分为载体接头序列。Underlined sequences are the parts of carrier connectors.
Table 1 表1 表1本研究中所使用的引物序列 Table 1Primer sequences used in this study
增PeU3-1和PeU3-2启动子全长。扩增体系及反应程序参照Phanta EVO HS Super-Fidelity DNA Polymerase (Cat No.P504)说明书。扩增产物经凝胶电泳检测后回收、纯化目的片段, 然后进行连接, 转化大肠杆菌感受态细胞。利用载体通用引物M13F/R进行菌液PCR和测序。
(A) 不同截短PeU3启动子驱动GUS在烟草叶片中的瞬时表达(Bars=5 mm); (B) 不同截短PeU3启动子驱动GUS在麻竹愈伤组织中的表达(Bars=5 mm)。CK: 阴性对照 Figure 4 GUS expression driven by different truncated PeU3 promoters
(A) Transient expression of GUS gene driven by different truncated PeU3 promoters in the leaves of Nicotiana benthamiana (Bars=5 mm); (B) Expression of GUS gene driven by different truncated PeU3 promoters in the callus of Dendrocalamus latiflorus (Bars=5 mm). CK: Negative control
(A), (B) PeU3启动子在烟草叶片中的瞬时表达LUC荧光图; (C) PeU3启动子在烟草叶片中的瞬时表达LUC相对活性(每个实验均设4个生物学重复。* 表示Duncan法检测时在P<0.05水平差异显著); CPS: 荧光强度 Figure 5LUC expression driven by different truncated PeU3 promoters in the leaves of Nicotiana benthamiana
(A), (B) Transient expression LUC assays illustrating the activation of PeU3 promoter in leaves of N. benthamiana; (C) The quantification of the relative luminescence intensities (In each experiment, four biological replications were performed with quanti?cation. Asterisk above the bars denote significant differences determined by the Duncan’s test, P<0.05); CPS: Luminescence intensity
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... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
In vivo genome editing using a high-efficiency TALEN system 1 2012
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
Plant genome editing made easy: targeted mutagenesis in model and crop plants using the CRISPR/Cas system 1 2013
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
Enhancing gene targeting with designed zinc finger nucleases 1 2003
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
Overexpression of both flowering time genes AtSOC1 and SaFUL revealed huge influence onto plant habitus in poplar 1 2019
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
Efficient targeted mutagenesis in apple and first time edition of pear using the CRISPR- Cas9 system 1 2019
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Copper induced oxidative stresses, antioxidant responses and phytoremediation potential of Moso bamboo (Phyllostachys pubescens) 1 2015
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
Multiplex genome engineering using CRISPR/Cas systems 3 2013
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
Single and multiple gene knockouts by CRISPR-Cas9 in maize 1 2019
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
The CRISPR/Cas bacterial immune system cleaves bacteriophage and plasmid DNA 1 2010
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
Main regulatory pathways, key genes and microRNAs involved in flower formation and development of Moso bamboo (Phyllostachys edulis) 1 2017
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity 2 2012
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Activities and specificities of CRISPR/Cas9 and Cas12a nucleases for targeted mutagenesis in maize 1 2019
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
The association of hormone signaling genes, transcription and changes in shoot anatomy during Moso bamboo growth 1 2018
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
Varied transcriptional efficiencies of multiple Arabidopsis U6 small nuclear RNA genes 1 2007
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes 1 2017
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
Targeted mutagenesis in Zea mays using TALENs and the CRISPR/ Cas system 1 2014
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants 1 2015
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
Characterization of the U3 and U6 snRNA genes from wheat: U3 snRNA genes in monocot plants are transcribed by RNA polymerase III 1 1992
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Heat- shock-inducible CRISPR/Cas9 system generates heritable mutations in rice 1 2019
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease 1 2013
Site-specific manipulation of Arabidopsis loci using CRISPR-Cas9 SunTag systems 1 2019
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
The draft genome of the fast-growing non-timber forest species Moso bamboo (Phyllostachys heterocycla) 1 2013
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
Recovery of the non-functional EGFP-assisted identification of mutants generated by CRISPR/Cas9 1 2019
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
In vitro propagation of bamboo species through axillary shoot proliferation: a review 1 2018
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
CRISPR/Cas9- mediated targeted mutagenesis in the liverwort Marchantia polymorpha L 1 2014
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
Hairpin RNAs derived from RNA polymerase II and polymerase III promoter-directed transgenes are processed differently in plants 1 2008
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
Robust CRISPR/Cas9 mediated genome editing and its application in manipulating plant height in the first generation of hexaploid Ma bamboo (Dendrocalamus latiflorus Munro) 3 2020
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
... 具有明确转录起始位点的U3和U6启动子是CRISPR/Cas9技术中驱动sgRNA转录的重要元件; 其转录起始位点分别为A和G, 且转录活性较高.选择具有明确起始位点的U3或U6启动子, 能精确引导sgRNA的转录从而减少无关DNA转录带来的脱靶效应(Li et al., 2007; Belhaj et al., 2013; Cong et al., 2013).虽然U3和U6启动子已在多个物种基因组编辑系统中得到应用, 但相同的U3或U6启动子在同源关系较远的物种间并不一定适用(Wang et al., 2008; Sugano et al., 2014).而且同一物种基因中往往存在多个U3和U6启动子, 其转录效率也不相同, 有些多位点基因编辑载体的构建需要多种U3和U6启动子.因此, 克隆出更多的植物内源U3/U6启动子, 有利于CRISPR/Cas9基因组编辑系统的完善.Ye等(2020)利用麻竹原生质体系统比较了3个水稻U6启动子(OsU6a/OsU6b/OsU6c)之间的编辑效率, 发现OsU6的编辑效率更高, 那么利用竹子内源U3/U6启动子是否会带来更精准且高效的编辑? 鉴于此, 本研究从毛竹中克隆了2种U3启动子, 并将其进行不同长度的截短, 构建相应的GUS及LUC表达载体, 利用农杆菌介导转化法侵染麻竹愈伤组织及烟草叶片, 通过GUS染色、活体成像及LUC活性测定, 验证不同截短U3启动子的转录活性, 以期为构建竹子CRISPR/ Cas9基因组编辑体系提供更多理想的内源启动子. ...
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Generation of new glutinous rice by CRISPR/Cas9-targeted mutagenesis of the Waxy gene in elite rice varieties 1 2018
... 优势, 与导向RNA (single guide RNA)结合的Cas9蛋白不仅能发挥识别的作用, 还可以行使切割功能, 靶向特定位点, 操作简单高效, 还可针对多个靶点同时进行编辑(Garneau et al., 2010; Jinek et al., 2012; Cong et al., 2013; Ma et al., 2015).目前, CRISPR/ Cas9系统作为一种高效的基因定点编辑技术, 已成功应用于拟南芥(Arabidopsis thaliana)、水稻(Oryza sativa)、玉米(Zea mays)、小麦(Triticum aestivum)、烟草(Nicotiana benthamiana)以及杨树(Populus simonii)等植物(Liang et al., 2017; Lee et al., 2018; Zhang et al., 2018; Bruegmann and Fladung, 2019; Papikian et al., 2019; Ren et al., 2019); 在竹子中也首次实现了对麻竹(Dendrocalamus latiflorus)八氢番茄红素合酶(phytoene synthase)基因PSY1的成功编辑(Ye et al., 2020). ...
Development of an Agrobacterium- delivered CRISPR/Cas9 system for wheat genome editing 1 2019
... CRISPR/Cas9系统最初在细菌中发现, 该系统对靶DNA进行切割时仅需1个Cas蛋白和2个RNA元件即可实现(Jinek et al., 2012).该技术因操作简单、突变率高、成本低等优势而快速发展, 为基因功能研究及作物遗传改良做出了巨大贡献.利用U3或U6启动子建立CRISPR/Cas9的基因组编辑系统已应用于多种植物(Charrier et al., 2019; Doll, 2019; Nandy et al., 2019; Zhang et al., 2019).虽然Ye等(2020)利用水稻U6启动子建立了麻竹基因组编辑体系, 但目前关于竹子内源U3/U6启动子的研究还未见报道.因此, 本研究从毛竹中克隆了2种PeU3启动子, 分析发现其含有比较保守的USE元件和TATA框, 与水稻的OsU3启动子类似.在玉米、番茄(Lycopersicon esculentum)及小麦的U3启动子中也发现了这2个保守元件(Marshallsay et al., 1992; 李丽莉等, 2005; Liang et al., 2014), 这表明本研究克隆出的2个毛竹PeU3启动子可能具有转录活性. ...
Chromosome-level reference genome and alternative splicing atlas of Moso bamboo (Phyllostachys edulis) 1 2018
... 竹子是重要的森林资源之一, 具有分布广、生长快、用途多、生态和经济价值高等特点.毛竹(Phyllostachys edulis)基因组测序的完成极大地推动了竹类植物分子生物学的发展(Peng et al., 2013; Zhao et al., 2018).近年来, 研究人员先后在竹子鞭芽发育、快速生长、开花及抗逆等方面开展了相关研究(Chen et al., 2015; Ge et al., 2017; Li et al., 2018; Sandhu et al., 2018; 孙建飞等, 2018); 但由于竹子分子生物学的许多研究技术体系还不够成熟, 大量与竹子重要经济性状相关的基因资源还未得到充分研究和利用.基因组编辑技术是一种新近发展起来的对基因组进行定向精确修饰的技术, 主要包括人工核酸酶介导的锌指核酸酶(zinc-finger nucleases, ZFN)、转录激活样效应因子核酸酶(transcription activator- like effector nucleases, TALEN)及RNA引导的CRISPR/Cas核酸酶(clustered regularly interspaced short palindromic repeat, CRISPR)技术(Bibikova et al., 2003; Bedell et al., 2012; Cong et al., 2013).基因组编辑技术在基础研究和作物遗传改良等方面展示出巨大的潜力, 其中第3代基因组编辑技术(如CRISPR/Cas9)拥有其它基因组编辑技术无可比拟的 ...
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新窗口打开|下载原图ZIP|生成PPT
新窗口打开|下载原图ZIP|生成PPT
新窗口打开|下载原图ZIP|生成PPT
新窗口打开|下载原图ZIP|生成PPT
新窗口打开|下载原图ZIP|生成PPT
