Tobacco (Nicotiana tabacum L.) seed lipid is a promising non-edible feedstock for biodiesel production. In order to meet the increasing demand, achieving high seed lipid content is one of the major goals in tobacco seed production. The TT8 gene and its homologs negatively regulate seed lipid accumulation in Arabidopsis and Brassica species. We speculated that manipulating the homolog genes of TT8 in tobacco could enhance the accumulation of seed lipid. In this present study, we found that the TT8 homolog genes in tobacco, NtAn1a and NtAn1b, were highly expressed in developing seed. Targeted mutagenesis of NtAn1 genes was created by the CRISPR-Cas9-based gene editing technology. Due to the defect of proanthocyanidin (PA) biosynthesis, mutant seeds showed the phenotype of a yellow seed coat. Seed lipid accumulation was enhanced by about 18 and 15% in two targeted mutant lines. Protein content was also significantly increased in mutant seeds. In addition, the seed yield-related traits were not affected by the targeted mutagenesis of NtAn1 genes. Thus, the overall lipid productivity of the NtAn1 knockout mutants was dramatically enhanced. The results in this present paper indicated that tobacco NtAn1 genes regulate both PAs and lipid accumulation in the process of seed development and that targeted mutagenesis of NtAn1 genes could generate a yellow-seeded tobacco variety with high lipid and protein content. Furthermore, the present results revealed that the CRISPR-Cas9 system could be employed in tobacco seed de novo domestication for biodiesel feedstock production.

烟草（烟草L.）种子脂质是用于生物柴油生产的有前途的不可食用原料。为了满足日益增长的需求，实现高种子脂质含量是烟草种子生产的主要目标之一。的TT8 基因及其同源物负调控大豆种子脂质积累 拟南芥 和 芸苔属种类。我们推测操纵基因的同源基因TT8烟草中的维生素可以增强种子脂质的积累。在本研究中，我们发现TT8 烟草中的同源基因 NtAn1a 和 NtAn1b，在发育中的种子中高度表达。靶向诱变NtAn1基因是通过基于CRISPR-Cas9的基因编辑技术创建的。由于原花青素（PA）生物合成的缺陷，突变种子显示出黄色种皮的表型。在两个靶向突变株中，种子脂质的积累提高了约18％和15％。突变种子中的蛋白质含量也显着增加。此外，与种子产量相关的性状不受定向诱变的影响。NtAn1基因。因此，NtAn1敲除突变体大大增强。本文的结果表明，烟草NtAn1 基因在种子发育过程中调节PAs和脂质积累，并靶向诱变。 NtAn1该基因可以产生具有高脂质和蛋白质含量的黄色种子烟草品种。此外，目前的结果表明，CRISPR-Cas9系统可用于烟草种子。从头 生物柴油原料的驯化。