CRISPR/Cas9 mediated knockout of the abdominal-A homeotic gene in the global pest, diamondback moth (Plutella xylostella).
The diamondback moth, Plutella xylostella (L.), is a worldwide agricultural pest that has developed resistance to multiple classes of insecticides. Genetics-based approaches show promise as alternative pest management approaches but require functional studies to identify suitable gene targets. Here we use the CRISPR/Cas9 system to target a gene, abdominal-A, which has an important role in determining the identity and functionality of abdominal segments. We report that P. xylostella abdominal-A (Pxabd-A) has two structurally-similar splice isoforms (A and B) that differ only in the length of exon II, with 15 additional nucleotides in isoform A. Pxabd-A transcripts were detected in all developmental stages, and particularly in pupae and adults. CRISPR/Cas9-based mutagenesis of Pxabd-A exon I produced 91% chimeric mutants following injection of 448 eggs. Phenotypes with abnormal prolegs and malformed segments were visible in hatched larvae and unhatched embryos, and various defects were inherited by the next generation (G1). Genotyping of mutants demonstrated several mutations at the Pxabd-A genomic locus. The results indicate that a series of insertions and deletions were induced in the Pxabd-A locus, not only in G0 survivors but also in G1 individuals, and this provides a foundation for genome editing. Our study demonstrates the utility of the CRISPR/Cas9 system for targeting genes in an agricultural pest and therefore provides a foundation the development of novel pest management tools.
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China.
Yuping
Y
Huang
Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
Yazhou
Y
Chen
Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
Baosheng
B
Zeng
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China.
Yajun
Y
Wang
Department of Microbiology & Molecular Genetics, University of California, Irvine, CA 92697-4025, USA; Department of Molecular Biology & Biochemistry, University of California, Irvine, CA 92697-3900, USA.
Anthony A
AA
James
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China; Graham Centre, Charles Sturt University, Orange, NSW 2800, Australia.
Geoff M
GM
Gurr
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China.
Guang
G
Yang
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China.
Xijian
X
Lin
Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China. Electronic address: yphuang@sibs.ac.cn.
Yongping
Y
Huang
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China; Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China. Electronic address: msyou@iae.fjau.edu.cn.
Minsheng
M
You
D000005:Abdomen / Q000254:growth & development / Q000502:physiology; D000818:Animals; D001483:Base Sequence; D064113:CRISPR-Cas Systems; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005801:Genes, Homeobox; D007814:Larva / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D008297:Male; D009036:Moths / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D010063:Ovum / Q000254:growth & development / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D029721:Drosophila Proteins; D009687:Nuclear Proteins; D014157:Transcription Factors; C066591:abd-A protein, Drosophila
Gene mutation; Germline stability; Lepidoptera; Pest; Phylogenetic analysis