In-situ Hybridization to Messenger RNA in Heterodera glycines
A method is presented for in-situ hybridization to mRNA in second-stage juveniles (J2) of the soybean cyst nematode Heterodera glycines. The protocol was developed using a digoxigenin-labeled RNA probe transcribed from cDNA of a cellulase gene that was known to be expressed in the subventral esophag...
| Publié dans: | Journal of nematology. - 1969. - 30(1998), 3 vom: 09. Sept., Seite 309-12 |
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| Auteur principal: | |
| Autres auteurs: | , , , |
| Format: | Article |
| Langue: | English |
| Publié: |
1998
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| Accès à la collection: | Journal of nematology |
| Sujets: | Journal Article Heterodera glycines cellulase gene digoxigenin RNA probe esophageal gland in-situ hybridization nematode |
| Résumé: | A method is presented for in-situ hybridization to mRNA in second-stage juveniles (J2) of the soybean cyst nematode Heterodera glycines. The protocol was developed using a digoxigenin-labeled RNA probe transcribed from cDNA of a cellulase gene that was known to be expressed in the subventral esophageal glands of H. glycines. Formaldehyde-fixed J2 were cut into sections with a vibrating razor blade to make the inside of the nematodes accessible for probing. These nematode fragments then were hybridized in suspension with riboprobe, and labeled with an alkaline phosphatase-conjugated antibody to digoxigenin. Staining with nitroblue tetrazolium and bromo-chloro-indolyl phosphate revealed a highly specific hybridization signal to mRNA within the cytoplasm of the subventral gland cells, using this specific antisense probe. This in-situ hybridization protocol will be useful for the characterization and identification of esophageal gland secretion genes in plant-parasitic nematodes, among other applications |
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| Description: | Date Completed 14.07.2011 Date Revised 20.10.2021 published: Print Citation Status PubMed-not-MEDLINE |
| ISSN: | 0022-300X |