RNA-interference methods for gene-knockdown in the sea louse, Lepeophtheirus salmonis

studies on a putative prostaglandin E synthase

Ewan M. Campbell, C. C. Pert, Alan S. Bowman (Corresponding Author)

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Harnessing the full utility of extensive gene sequences recently available for the economically important sea louse Lepeophtheirus salmonis, requires the adaptation of modern molecular biology approaches to this non-model organism. Using a putative microsomal prostaglandin E synthase type-2 (PGES2) as a candidate gene, we investigated gene- knockdown by double-stranded RNA interference (dsRNAi) in the small free-living and the larger parasitic stages of L. salmonis. dsRNA was administered to nauplis and copepodid stages by immersion for 7 h. Pre-adult and adults received dsRNA by intra-haemocoelic injection. The extent, speed mid persistence of the knockdown effects were determined by RT-PCR. LsPGES2 was abundantly expressed in all life stages, including the non-parasitic stages. Administration of dsRNA to nauplius and copepodids by immersion had no effect on mortality rates and moulting through to copepodids was observed. Dramatic knockdown of LsPGES2 was observed within 7 h and persisted for at least 48 h. Injection of dsRNA lipid no effect on mortality in pre-adults and adults, but knockdown of LsPGES2 was apparent within 24 h reaching 95% over the 72 h and was persisterit for at least 120 h. The methods developed resulted ill rapid mid persistent knockdown ill L. salmonis suitable for studies in the different stadia.

Original languageEnglish
Pages (from-to)867-874
Number of pages8
JournalParasitology
Volume136
Issue number8
DOIs
Publication statusPublished - Jul 2009

Keywords

  • sea lice
  • Lepeophtheirus salmonis
  • gene knockdown
  • RNA interference
  • dsRNAi
  • microsomal prostaglandin E synthase
  • MAPEG
  • glutathione transferase
  • products
  • industry
  • target

Cite this

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title = "RNA-interference methods for gene-knockdown in the sea louse, Lepeophtheirus salmonis: studies on a putative prostaglandin E synthase",
abstract = "Harnessing the full utility of extensive gene sequences recently available for the economically important sea louse Lepeophtheirus salmonis, requires the adaptation of modern molecular biology approaches to this non-model organism. Using a putative microsomal prostaglandin E synthase type-2 (PGES2) as a candidate gene, we investigated gene- knockdown by double-stranded RNA interference (dsRNAi) in the small free-living and the larger parasitic stages of L. salmonis. dsRNA was administered to nauplis and copepodid stages by immersion for 7 h. Pre-adult and adults received dsRNA by intra-haemocoelic injection. The extent, speed mid persistence of the knockdown effects were determined by RT-PCR. LsPGES2 was abundantly expressed in all life stages, including the non-parasitic stages. Administration of dsRNA to nauplius and copepodids by immersion had no effect on mortality rates and moulting through to copepodids was observed. Dramatic knockdown of LsPGES2 was observed within 7 h and persisted for at least 48 h. Injection of dsRNA lipid no effect on mortality in pre-adults and adults, but knockdown of LsPGES2 was apparent within 24 h reaching 95{\%} over the 72 h and was persisterit for at least 120 h. The methods developed resulted ill rapid mid persistent knockdown ill L. salmonis suitable for studies in the different stadia.",
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T1 - RNA-interference methods for gene-knockdown in the sea louse, Lepeophtheirus salmonis

T2 - studies on a putative prostaglandin E synthase

AU - Campbell, Ewan M.

AU - Pert, C. C.

AU - Bowman, Alan S.

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N2 - Harnessing the full utility of extensive gene sequences recently available for the economically important sea louse Lepeophtheirus salmonis, requires the adaptation of modern molecular biology approaches to this non-model organism. Using a putative microsomal prostaglandin E synthase type-2 (PGES2) as a candidate gene, we investigated gene- knockdown by double-stranded RNA interference (dsRNAi) in the small free-living and the larger parasitic stages of L. salmonis. dsRNA was administered to nauplis and copepodid stages by immersion for 7 h. Pre-adult and adults received dsRNA by intra-haemocoelic injection. The extent, speed mid persistence of the knockdown effects were determined by RT-PCR. LsPGES2 was abundantly expressed in all life stages, including the non-parasitic stages. Administration of dsRNA to nauplius and copepodids by immersion had no effect on mortality rates and moulting through to copepodids was observed. Dramatic knockdown of LsPGES2 was observed within 7 h and persisted for at least 48 h. Injection of dsRNA lipid no effect on mortality in pre-adults and adults, but knockdown of LsPGES2 was apparent within 24 h reaching 95% over the 72 h and was persisterit for at least 120 h. The methods developed resulted ill rapid mid persistent knockdown ill L. salmonis suitable for studies in the different stadia.

AB - Harnessing the full utility of extensive gene sequences recently available for the economically important sea louse Lepeophtheirus salmonis, requires the adaptation of modern molecular biology approaches to this non-model organism. Using a putative microsomal prostaglandin E synthase type-2 (PGES2) as a candidate gene, we investigated gene- knockdown by double-stranded RNA interference (dsRNAi) in the small free-living and the larger parasitic stages of L. salmonis. dsRNA was administered to nauplis and copepodid stages by immersion for 7 h. Pre-adult and adults received dsRNA by intra-haemocoelic injection. The extent, speed mid persistence of the knockdown effects were determined by RT-PCR. LsPGES2 was abundantly expressed in all life stages, including the non-parasitic stages. Administration of dsRNA to nauplius and copepodids by immersion had no effect on mortality rates and moulting through to copepodids was observed. Dramatic knockdown of LsPGES2 was observed within 7 h and persisted for at least 48 h. Injection of dsRNA lipid no effect on mortality in pre-adults and adults, but knockdown of LsPGES2 was apparent within 24 h reaching 95% over the 72 h and was persisterit for at least 120 h. The methods developed resulted ill rapid mid persistent knockdown ill L. salmonis suitable for studies in the different stadia.

KW - sea lice

KW - Lepeophtheirus salmonis

KW - gene knockdown

KW - RNA interference

KW - dsRNAi

KW - microsomal prostaglandin E synthase

KW - MAPEG

KW - glutathione transferase

KW - products

KW - industry

KW - target

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DO - 10.1017/S0031182009990357

M3 - Article

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SP - 867

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JO - Parasitology

JF - Parasitology

SN - 0031-1820

IS - 8

ER -