在Populus中基于CRISPR/Cas9的基因激活和基因编辑
Tao Yao1,2, Guoliang Yuan1,2,3, Haiwei Lu1,4
1Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA.
Horticulture research
|June 16, 2023
概括
这项研究表明了树中CRISPR激活和基编辑技术的有效性. 这些先进的基因组编辑工具能够在木质物种中精确调节基因表达和基因工程,以改善特征.
科学领域:
- 植物生物技术 植物生物技术
- 分子生物学分子生物学
- 森林遗传学 森林遗传学
背景情况:
- 种类*Populus*对于环境,工业和生物燃料应用至关重要,作为树木研究的典范.
- 克里斯普尔/Cas9已被用于*Populus*中的基因淘汰,但基因激活和基因编辑技术在很大程度上仍未得到评估.
- 对Populus的遗传改进对于增强生长和量身定制的红素成分至关重要.
研究的目的:
- 为了评估非活性化Cas9 (dCas9) 基CRISPR激活 (CRISPRa) 在两个*Populus*克隆中对基因表达微调的有效性.
- 评估基于Cas9尼克酶 (nCas9) 的细胞酶基编辑器 (CBE) 在*Populus*中用于精确基因工程的应用.
- 为了证明基于CRISPR/Cas的先进技术在木质物种的基因调节和工程方面的实用性.
主要方法:
- 采用基于dCas9的CRISPRa技术调节*TPX2*和*LecRLK-G*基因的表达.
- 利用原生质中的短暂表达和*Agrobacterium*介导的稳定转化用于CRISPRa传递.
- 应用基于nCas9的CBE在*PLATZ*基因中引入C-to-T转换,用于过早停止密码子诱导.
主要成果:
- 在短暂和稳定系统中,CRISPRa成功地将目标基因表达增加了1.2倍至7.0倍.
- 基于Cas9尼克酶的细胞酶基编辑器在*PLATZ*基因中引入C-to-T突变方面实现了13-14%的效率.
- 证明了基于dCas9的CRISPRa和基于nCas9的CBE在*Populus*物种中的有效性.
结论:
- 成功地应用了基于CRISPR/Cas的技术来调节基因表达和精确的基因工程在两个*Populus*物种.
- 基于dCas9的CRISPRa系统是有效的微调基因表达在*Populus*.
- 像CRISPRa和CBE这样的新兴基因组编辑工具可以很容易地被采用来推进木质物种的研究和应用.
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