公开(公告)号：EP3155001A1

公开(公告)日：2017-04-19

申请号：EP15730110.2

申请日：2015-06-12

申请人： SESVanderHave N.V.

发明人： WEYENS, Guy ,  LEFÈBVRE, Marc ,  THAMMANNAGOWDA, Shivegowda ,  LOMMEL, Murielle ,  SONG, Luo

IPC分类号： C07K14/415 ,  C12N15/82

CPC分类号： C12N15/8213 ,  C12N9/88

摘要： A method for transformation of sugar beet protoplasts includes obtaining protoplasts from stomatal guard cells isolated from a sugar beet plant. The protoplasts are transformed with a nucleic acid construct including a nucleotide sequence of interest and Transcription Activator-Like Effector Nucleases (TALEN) or one or more vectors including sequences encoding these Transcription Activator-Like Effector Nucleases (TALEN)sequences. The TALEN target and process a target sequence and replace the target sequence through homologous recombination with the nucleic acid construct including the nucleotide sequence of interest, -possibly applying to an in vitroculture of the protoplasts, a medium that is toxic, preferably lethal to the in vitroculture of the protoplasts. Sugar beet plants are regenerated from the cell culture, preferably from the surviving protoplasts having integrated the nucleic acid construct including the sequence of interest that possibly renders the transformed cell resistant to the toxic activity of the applied medium.

摘要翻译： 用于转化甜菜原生质体的方法包括以下步骤： - 从由甜菜植物分离的气孔保卫细胞获得原生质体， - 用包含目的核苷酸序列和转录激活物样效应物核酸酶的核酸构建体转化所述原生质体（ TALEN）或包含编码这些转录激活物样效应物核酸酶（TALEN）序列的序列的一种或多种载体，其中这些TALEN靶向并加工靶序列并通过与包含目标核苷酸序列的核酸构建体同源重组来替换该靶序列 ，可能适用于这些原生质体的培养物，有毒的，优选对原生质体的培养物致死的培养基，以及 - 从细胞培养物中再生甜菜植物，优选来自已整合有核酸构建体的存活原生质体 包括可能的兴趣序列 使得转化的细胞对所施用的培养基的毒性具有抗性。

公开(公告)号：EP3155000A1

公开(公告)日：2017-04-19

申请号：EP15730106.0

申请日：2015-06-12

申请人： SESVanderHave N.V.

发明人： WEYENS, Guy ,  LEFÈBVRE, Marc ,  THAMMANNAGOWDA, Shivegowda ,  LOMMEL, Murielle

IPC分类号： C07K14/415 ,  C12N15/82

CPC分类号： C12N15/821 ,  A01H1/02 ,  A01H1/04 ,  C12N9/88 ,  C12N15/52 ,  C12N15/8209 ,  C12N15/8241 ,  C12N15/8279 ,  C12N15/8281 ,  C12N15/8282 ,  C12N15/8285 ,  C12N15/8286

摘要： A method for the transformation of sugar beet protoplasts includes obtaining protoplasts from stomatal guard cells isolated from a sugar beet plant. The protoplasts are transformed with a nucleic acid construct including a nucleotide sequence of interest and a selection marker sequence. One or more ALS inhibitors at a concentration that is lethal to the in vitro culture of the protoplasts are applied to an in vitro culture of the protoplasts. Sugar beet plants are regenerated from the surviving protoplasts having integrated the nucleic acid construct including the sequence of interest and the selection marker sequence. The selection marker sequence is the mutated BvALS113 sequence carrying in its sequence a mutation at amino acid 113 position from Alanine to Tyrosine.

摘要翻译： 本发明涉及一种用于转化甜菜原生质体的方法，包括以下步骤： - 从分离自甜菜植物的气孔保护细胞获得原生质体， - 用包含感兴趣的核苷酸序列的核酸构建体转化原生质体，以及 选择标记序列， - 应用于所述原生质体的体外培养，一种或多种浓度对原生质体的体外培养致死的ALS抑制剂，以及从具有整合核酸的存活的原生质体再生甜菜植物 构建体，其包含感兴趣的序列和选择标记序列，其中选择标记序列是突变的BvALS113序列，其在氨基酸113位置从丙氨酸到酪氨酸的突变。