title: Creation of fragrant rice by targeted knockout of the OsBADH2 gene using TALEN technology.
abstract: Fragrant rice is favoured worldwide because of its agreeable scent. The presence of a defective badh2 allele encoding betaine aldehyde dehydrogenase (BADH2) results in the synthesis of 2-acetyl-1-pyrroline (2AP), which is a major fragrance compound. Here, transcription activator-like effector nucleases (TALENs) were engineered to target and disrupt the OsBADH2 gene. Six heterozygous mutants (30%) were recovered from 20 transgenic hygromycin-resistant lines. Sanger sequencing confirmed that these lines had various indel mutations at the TALEN target site. All six transmitted the BADH2 mutations to the T1 generation; and four T1 mutant lines tested also efficiently transmitted the mutations to the T2 generation. Mutant plants carrying only the desired DNA sequence change but not the TALEN transgene were obtained by segregation in the T1 and T2 generations. The 2AP content of rice grains of the T1 lines with homozygous mutations increased from 0 to 0.35-0.75 mg/kg, which was similar to the content of a positive control variety harbouring the badh2-E7 mutation. We also simultaneously introduced three different pairs of TALENs targeting three separate rice genes into rice cells by bombardment and obtained lines with mutations in one, two and all three genes. These results indicate that targeted mutagenesis using TALENs is a useful approach to creating important agronomic traits.
authors.affiliation: State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Qiwei
authors.initials: Q
authors.lastname: Shan
authors.affiliation: State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Kunling
authors.initials: K
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.|Beijing Genovo Biotechnology Co. Ltd, Beijing, China.
authors.forename: Kang
authors.initials: K
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Caixia
authors.initials: C
authors.lastname: Gao
mesh_terms: D001483:Base Sequence; D050728:Betaine-Aldehyde Dehydrogenase / Q000235:genetics*; D018744:DNA, Plant; D055785:Gene Knockdown Techniques; D017343:Genes, Plant; D008969:Molecular Sequence Data; D009154:Mutation; D012275:Oryza / Q000201:enzymology / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
chemical_list: D018744:DNA, Plant; D050728:Betaine-Aldehyde Dehydrogenase
keywords: 2AP; betaine aldehyde dehydrogenase; genome editing; multiplex gene knockout; transcription activator-like effector nucleases