Topic: Single Cell Expression Analysis of Stem Cell Differentiation
Speaker:   Dr. Richard Fekete, Senior Manager of Life Technologies
Date: 29 November 2012 (Thu)
Time: 11:00 am -12:00 pm
Venue: Room 4020, 4/F, Lift 2, Academic Building, The Hong Kong University of Science & Technology, Clearwater Bay

Abstract

Human Embryonic Stem Cells (hESC) can be differentiated into primary progenitor cells which then in turn differentiate into a variety of functionally specialized cell types. One example of this is the differentiation of hESC to Neural Stem Cells (NSC), which then can differentiate into neurons, oligodendrocytes or astrocytes. A more thorough understanding of the differentiation pathway and associated factors are required. The pathway for differentiation is not a uniform process for every cell involved – some cells differentiate earlier than others and at different rates. Using FACS we sort thousands of single cells at different time points during differentiation, stabilize their profiles, preamplify 100 selected genes and analyze these by qPCR. In this way, profiles for numerous cells that have undergone the hESC to NSC pathway are defined, which will in turn enable an in-depth analysis of the growth factor dependence of NSCs.

All are welcome but limited seats will be allocated on first-come, first-served basis.
Questions? Please contact our Marketing team at 31077600 or email at hkmarketing@lifetech.com

Join the technical seminar
Date: 29 November 2012 (Thu)
Venue: Room 7-03, 7/F, The Hong Kong Jockey Club Building for Interdisciplinary Research, 5 Sassoon Road, Pokfulam, Hong Kong
Speaker:   Dr. Richard Fekete, Senior Manager of Life Technologies
Time Topics
3:00 - 4:00 p.m. Successful RNA workflow – Ambion sample preparation & analysis tools
4:00 - 4:15 p.m. Coffee break
4:15 - 5:15 p.m. Single Cell Expression Analysis

Abstract

Successful RNA workflow – Ambion sample preparation & analysis tools
RNA analysis provides an abundance of gene expression information and involves many steps. Often overlooked, stabilizing samples is often the most common time for RNA degradation. Purification of high quality RNA and miRNA with efficient DNA removal is the second key step, and can be done using a variety of methods such as magnetic bead based, filter based or organic extraction. Lysate based preparation products enable a rapid method of getting samples ready for analysis without compromising results. Enriching for targets of interest, either through positive selection or negative selection is also important in obtaining quality data. Accurate, simple and automation friendly methods for qPCR or the construction small RNA or whole transcriptome RNA libraries for sequencing followed by data analysis are the final steps in the RNA analysis workflow. Experimental data will be presented to address each of these stages in the RNA Sample to Analysis workflow.

Single Cell Expression Analysis
When analyzing gene expression profiles from large numbers of cells, the average profile may not be a true representation of the many different profiles that could exist in the cell population (ex., in different states of growth, differentiation or activation). The transcriptional variability of individual cells and any insight into the relationship between specific genes gets lost. One aspect of this emerging field that still needs to be developed is data analysis. How to present data from single cell experiments? What are the proper controls and/or normalization methods to be used? Can you confidently identify sub-populations? The data analysis of high sample numbers with a reduced number of targets is not as straightforward as when using many targets with a few samples (i.e. arrays). Performing cluster analysis and displaying data as histograms can mask the identification of sub-populations. Using FACS we sort thousands of single cells at different time points during stem cell differentiation, stabilize their profiles, preamplify 100 selected genes and analyze these by qPCR. This gives the relative copy number for each transcript in a single cell and allows cells to be classified based on expression profiles. In this way, profiles for numerous cells that have undergone the hESC to NSC pathway are defined, which will in turn enable an in-depth analysis of the growth factor dependence of NSCs, the development of serum free media and the scale-up production of pure NSC from hESCs.

All are welcome but limited seats will be allocated on first-come, first-served basis.
Questions? Please contact our Marketing team at 31077600 or email at hkmarketing@lifetech.com

Join the technical seminar