This book applies modern molecular diagnostic techniques to the analysis of single cells, small numbers of cells, or cell extracts. Emphasis is placed on non-invasive analysis of single cell metabolites and the direct analysis of RNA and DNA from single cells, with a focus on polymerase chain reaction and fluorescence in situ hybridization.
The authors, all leading scientists in the field, provide current and innovative methods for extracting useful, accurate, and reliable data from single-cell samples. The stepwise, technically precise protocols presented in this volume thoroughly describe single-cell assays, from material collection and isolation of cells to data interpretation. Cell-types analyzed include embryonic blastomeres, fibroblasts, and lymphocytes. Among the techniques described are real-time quantitative PCR, isothermal whole genome amplification, comparative genomic hybridization, real-time genetic expression analysis, and the production of RNA and cDNA libraries.
This handbook is essential for researchers and clinicians, and practitioners providing care for couples seeking treatment for infertility or preimplantation genetic diagnosis.
Table of contents
Table of Contents
1. Non-invasive metabolic assessment of single cells
David K. Gardner
2. Laser Capture Microdissection for Analysis of Single Cells
Nobuki Nakamura, Katharina Ruebel, Long Jin, Xiang Qian, Heyu Zhang, and Ricardo V. Lloyd
3. Fluorescence in situ hybridization (FISH) on Single Cells (Sex Determination and Chromosome Rearrangements)
Paul N. Scriven and Caroline Mackie Ogilvie
4. Single cell PCR for PGD: methods, strategies and limitations
Karen Sermon and Martine De Rycke
5. Real-Time Quantitative PCR Measurement of Male Fetal DNA in maternal plasma
Bernhard Zimmermann, Ahmad El-Sheikhah, Kypros Nicolaides, Wolfgang Holzgreve, Sinuhe Hahn
6. Cell-Free Fetal DNA Plasma Extraction and Real Time-PCR Quantification
Jill L. Maron, Kirby L. Johnson, and Diana W. Bianchi
7. LATE-PCR and allied technologies: Real-time detection strategies for rapid, reliable diagnosis from single cells
Kenneth E. Pierce and Lawrence J. Wangh
8. Efficient isothermal amplification of the entire genome from single cells
Karen V. Schowalter, Jolene R. Fredrickson and Alan R Thornhill
9. Comparative Genomic Hybridization on single cells
Lucille Voullaire and Leeanda Wilton
10. Generation of amplified RNAs and cDNA libraries from single mammalian cells
James Adjaye
11. Real-time PCR to measure gene expression in single cells
Dagan Wells
12. Gender Determination and Detection of Aneuploidy in Single Cells Using DNA Array-Based Comparative Genomic Hybridisation (Array CGH)
Dong Gui Hui, Xin Yuan Guan, and Nicole Hussey
13. Microarray technology for mutation analysis of low template DNA samples
Chelsea Salvado and David Cram