Procedure for rapid oocyte selection based on quantitative analysis of cumulus cell gene expression

Purpose To develop a procedure for the analysis of gene expression in cumulus cells during the interval between ovum pick up and insemination to select the best oocytes for fertilization. Methods Five RNA extraction methods, three reverse transcription procedures followed by Real-time quantitative P...

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Bibliographic Details
Published in:Journal of assisted reproduction and genetics 2010-07, Vol.27 (7), p.429-434
Main Authors: Ferrari, Stefania, Lattuada, Debora, Paffoni, Alessio, Brevini, Tiziana A. L., Scarduelli, Claudia, Bolis, Giorgio, Ragni, Guido, Gandolfi, Fulvio
Format: Article
Language:English
Subjects:
Cumulus Cells - metabolism
Embryos
Female
Fertilization in Vitro
Gene Expression
Gynecology
Human Genetics
Humans
Medicine
Medicine & Public Health
Oocyte Retrieval
Oocytes - physiology
Ovaries
Ovulation
Quantitative analysis
Reproductive Medicine
Reverse Transcriptase Polymerase Chain Reaction - methods
RNA, Messenger - genetics
RNA, Messenger - metabolism
Technical Innovations
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Summary:Purpose To develop a procedure for the analysis of gene expression in cumulus cells during the interval between ovum pick up and insemination to select the best oocytes for fertilization. Methods Five RNA extraction methods, three reverse transcription procedures followed by Real-time quantitative PCR and one single-step mRNA quantification kit were tested to measure the expression of five genes in cumulus cells. Results Two RNA extraction kits gave the best combination of efficiency and purity. One reverse transcription procedure gave the best speed and efficiency. The single-step kit required more biological material than would be available from single cumulus oocyte complexes (COCs). Conclusions Our test identified a combination of RNA extraction and reverse transcription procedures that enables the level measurement of 5 selected cumulus cell transcripts within 4 h. Using this combination it was possible to obtain a reliable quantification of gene expression in 44 out of 46 individual COCs collected from seven patients.
ISSN:1058-0468
1573-7330
DOI:10.1007/s10815-010-9428-1