Quantitative Detection of Double-Stranded RNA-Mediated Gene Silencing of Parasitism Genes in Heterodera glycines
The introduction of a double-stranded RNA (dsRNA) into an organism to induce sequence-specific RNA interference (RNAi) of a target transcript has become a powerful technique to investigate gene function in nematodes and many organisms. Data provided here indicate that the inclusion of 1-2 mM spermid...
| Publié dans: | Journal of nematology. - 1969. - 39(2007), 2 vom: 28. Juni, Seite 145-52 |
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| Auteur principal: | |
| Autres auteurs: | , , , , , , |
| Format: | Article |
| Langue: | English |
| Publié: |
2007
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| Accès à la collection: | Journal of nematology |
| Sujets: | Journal Article |
| Résumé: | The introduction of a double-stranded RNA (dsRNA) into an organism to induce sequence-specific RNA interference (RNAi) of a target transcript has become a powerful technique to investigate gene function in nematodes and many organisms. Data provided here indicate that the inclusion of 1-2 mM spermidine and 50 mM octopamine and a 24 hr incubation period of nematodes in double-stranded RNA (dsRNA) soaking solutions resulted in a considerable increase in the percentage of nematodes that ingested dsRNA as compared to previous reports. This modified dsRNA soaking method was coupled with quantitative real-time RT-PCR (qRT-PCR) analyses to assess the potential silencing of the Heterodera glycines parasitism gene transcripts Hg-pel-1 and Hg-4E02 that are expressed within the esophageal gland cells of preparasitic H. glycines J2. The Hg-pel-1 transcript was most efficiently silenced with one dsRNA construct (ds267) at the highest dsRNA soaking concentration of 5.0 mg/ml, while the Hg-4E02 transcript was more efficiently silenced at the 2.5 mg/ml dsRNA concentration as compared to 5.0 mg/ml. A dsRNA construct (ds285) complementary to a different sequence within the Hg-pel-1 transcript than construct ds267 induced only minimal silencing of the Hg-pel-1 transcript at 2.5 mg/ml. The results suggest that both dsRNA concentration and sequence relative to the transcript targeted are critical for maximizing potential RNAi effects in parasitic nematodes |
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| Description: | Date Completed 25.09.2012 Date Revised 16.03.2024 published: Print Citation Status PubMed-not-MEDLINE |
| ISSN: | 0022-300X |