Genome editing and targeted sequence insertion with CrispR/Cas9 to elevate disease resistance in potato and tomato (JONES_S17DTP)

CrispR/Cas9等可编程核酸酶通过使特定基因组序列成为突变或同源重组（HR）的靶标，正在彻底改变生物学。这种新育种技术（nbt）可以导致功能缺失突变，或者通过HR将一株植物的有用基因精确替换（Knock-In Exchange， KIE）到另一株植物的相应染色体位置。我们从野生马铃薯和番茄亲缘种中提取抗疫霉基因。本项目的主要目标是利用CrispR/Cas9（并测试相关的Cpf1可编程核酸酶）用功能性Rpi基因等位基因替换番茄和马铃薯二倍体自交系中无功能Rpi基因同源物。我们研究的是来自www.solynta.com的自交系土豆。在使用KIE靶向之前，将使用具有长PacBio reads的RenSeq对感兴趣的茄基因型的抗性基因库进行表征（Witek et al . 2016）。该学生将产生KIE事件（取决于NBT法规），可以在F1混合动力中推向市场，并将与私营和公共部门的育种者进行互动。Cermak T（2015）高频，精确修饰番茄基因组基因组生物学16:232 DOI 10.1186/s13059-015-0796-9 Witek (2016) Rpi基因克隆使用Renseq和sMrt测序Nat Biot 34:656-660 www.ncbi.nlm.nih.gov/pubmed/27111721

Programmable nucleases such as CrispR/Cas9 are revolutionizing biology by enabling specific genome sequences to be targeted for mutation, or for homologous recombination (HR). Such New Breeding Technologies (NBTs) can result in loss-of-function mutations, or enable precise replacement (Knock-In Exchange, or KIE) of useful genes from one plant via HR into the corresponding chromosomal position of another plant. We work with Resistance to Phytophthora infestans (Rpi) genes from wild potato and tomato relatives. The main goal of this project is to use CrispR/Cas9 (and test also the related Cpf1 programmable nuclease) to replace non-functional Rpi gene homologs in tomato and potato diploid inbred lines, with functional Rpi gene alleles. We work with inbred potato lines from www.solynta.com. Prior to targeting with KIE, resistance gene repertoires of Solanum genotypes of interest will be characterized using RenSeq with long PacBio reads (Witek et al 2016). The student will generate KIE events that (depending on NBT regulation) could be brought to market in F1 hybrids, and will interact with private and public sector breeders.Cermak T (2015) High-frequency, precise modification of the tomato genome Genome Biology 16:232 DOI 10.1186/s13059-015-0796-9 Witek (2016) Rpi gene cloning using Renseq and sMrt sequencing Nat Biot 34:656-660 www.ncbi.nlm.nih.gov/pubmed/27111721