Nicolia Alessandro, Proux-Wéra Estelle, Åhman Inger, Onkokesung Nawaporn, Andersson Mariette, Andreasson Erik, Zhu Li-Hua
Department of Plant Breeding, Swedish University of Agricultural Sciences, Alnarp, Sweden; Department of Plant Protection Biology, Swedish University of Agricultural Sciences, Alnarp, Sweden.
Department of Plant Protection Biology, Swedish University of Agricultural Sciences, Alnarp, Sweden.
J Biotechnol. 2015 Jun 20;204:17-24. doi: 10.1016/j.jbiotec.2015.03.021. Epub 2015 Apr 4.
Potato is the third largest food crop in the world, however, the high degree of heterozygosity, the tetrasomic inheritance and severe inbreeding depression are major difficulties for conventional potato breeding. The rapid development of modern breeding methods offers new possibilities to enhance breeding efficiency and precise improvement of desirable traits. New site-directed mutagenesis techniques that can directly edit the target genes without any integration of recombinant DNA are especially favorable. Here we present a successful pipeline for site-directed mutagenesis in tetraploid potato through transient TALEN expression in protoplasts. The transfection efficiency of protoplasts was 38-39% and the site-directed mutation frequency was 7-8% with a few base deletions as the predominant type of mutation. Among the protoplast-derived calli, 11-13% showed mutations and a similar frequency (10%) was observed in the regenerated shoots. Our results indicate that the site-directed mutagenesis technology could be used as a new breeding method in potato as well as for functional analysis of important genes to promote sustainable potato production.
马铃薯是世界上第三大粮食作物，然而，高度杂合性、四体遗传和严重的近亲繁殖衰退是传统马铃薯育种的主要困难。现代育种方法的快速发展为提高育种效率和精准改良优良性状提供了新的可能性。能够直接编辑目标基因而无需任何重组DNA整合的新型定点诱变技术尤其有利。在此，我们展示了一种通过在原生质体中瞬时表达TALEN在四倍体马铃薯中进行定点诱变的成功流程。原生质体的转染效率为38 - 39%，定点突变频率为7 - 8%，少数碱基缺失是主要的突变类型。在原生质体来源的愈伤组织中，11 - 13%表现出突变，在再生芽中观察到类似的频率（10%）。我们的结果表明，定点诱变技术可作为马铃薯的一种新育种方法以及用于重要基因的功能分析，以促进马铃薯的可持续生产。
