title: Parallel WGA and WTA for Comparative Genome and Transcriptome NGS Analysis Using Tiny Cell Numbers.
abstract: Genomic DNA determines how and when the transcriptome is changed by a trigger or environmental change and how cellular metabolism is influenced. Comparative genome and transcriptome analysis of the same cell sample links a defined genome with all changes in the bases, structure, or numbers of the transcriptome. However, comparative genome and transcriptome analysis using next-generation sequencing (NGS) or real-time PCR is often limited by the small amount of sample available. In mammals, the amount of DNA and RNA in a single cell is ∼10 picograms, but deep analysis of the genome and transcriptome currently requires several hundred nanograms of nucleic acids for library preparation for NGS sequencing. Consequently, accurate whole-genome amplification (WGA) and whole-transcriptome amplification (WTA) is required for such quantitative analysis. This unit describes how the genome and the transcriptome of a tiny number of cells can be amplified in a highly parallel and comparable process. Protocols for quality control of amplified DNA and application of amplified DNA for NGS are included.
authors.affiliation: Qiagen, Hilden, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Korfhage
authors.affiliation: Qiagen, Hilden, Germany.
authors.forename: Evelyn
authors.initials: E
authors.lastname: Fricke
authors.affiliation: Qiagen, Hilden, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Meier
mesh_terms: D000818:Animals; D003584:Cytological Techniques / Q000379:methods*; D020869:Gene Expression Profiling / Q000379:methods*; D023281:Genomics / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D008322:Mammals; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
keywords: cancer; genome analysis; next-generation sequencing (NGS); somatic genome variation; transcriptome analysis; whole-genome amplification (WGA); whole-transcriptome amplification (WTA)