Analysis of chitin synthase function in a plant parasitic nematode, Meloidogyne artiellia, using RNAi.
A single chitin synthase gene is responsible for chitin production in the eggshells of the plant parasitic nematode Meloidogyne artiellia. In this paper we describe a functional analysis of this gene using RNAi as well as further analysis of two similar genes from the free-living nematode Caenorhabditis elegans. The parasitic life-style of M. artiellia required the development of a novel method for delivery of dsRNA to nematode eggs that may be of utility in other experimental systems. C. elegans chitin synthase genes were silenced by feeding nematodes bacteria expressing appropriate chitin synthase dsRNA from a plasmid vector, while M. artiellia egg masses were soaked in dsRNA solution. The results obtained demonstrated that the synthesis of chitin continues to take place in nematode eggs within the egg sac in the parasitic nematode, and that the removal of this activity affects egg development in both C. elegans and M. artiellia. The method described here provides a new way of investigating gene function in plant parasitic nematodes allowing the validity of potential target genes for novel control methods to be assessed. Furthermore, since intact egg cells within the gelatinous matrix of M. artiellia are permeable to dsRNA, eggs of other nematodes may also be similarly permeable to dsRNA and therefore amenable to use with dsRNAi.
Fanelli
Elena
E
Dipartimento di Biochimica e Biologia Molecolare, University of Bari, Via Orabona 4, 70126 Bari, Italy.
Di Vito
Mauro
M
Jones
John T
JT
De Giorgi
Carla
C
D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics; D002686:Chitin / Q000096:biosynthesis; D002687:Chitin Synthase / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D005260:Female; D017238:Genes, Helminth; D008018:Life Cycle Stages; D009348:Nematoda / Q000469:parasitology* / Q000502:physiology; D010063:Ovum / Q000187:drug effects / Q000378:metabolism; D010944:Plants / Q000469:parasitology*; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000494:pharmacology; D018510:RNA, Helminth / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D012996:Solutions; D014431:Tylenchoidea / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development
D012330:RNA, Double-Stranded; D018510:RNA, Helminth; D012996:Solutions; D002686:Chitin; D002687:Chitin Synthase