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Describing a Transcription Factor Dependent Regulation of the MicroRNA Transcriptome
Published on: June 15, 2016
Transcriptome analyses of CHO cells with the next-generation microarray CHO41K: development and validation by
Jennifer Becker1, Christina Timmermann1, Oliver Rupp2
1Centrum für Biotechnologie, Universität Bielefeld, 33594 Bielefeld, Germany; AG Zellkulturtechnik, Technische Fakultät, Universität Bielefeld, 33594 Bielefeld, Germany.
Abstract:
The increasing importance of Chinese hamster ovary (CHO) cells for the production of pharmaceutical proteins has awakened the demand to understand the cellular metabolism of these cells. However, satisfactory gene expression studies have yet been impractical due to insufficient coverage of sequences. In this work, previously determined sequence information of CHO cells and newly derived data from 454 and Illumina sequencing was used to establish the CHO41K microarray which contains 41,304 probes. Self-hybridisation was performed for replica determination and samples were run in triplicates to increase statistical power. For determination of technical variance, confidence intervals at an M-value of ±0.6 for 95% and at ±2.3 for 99% of the probes were calculated. Intra-microarray and slide to slide variance was not detectable. In a first application, this microarray enabled an in-depth look inside the cellular transcriptome of CHO cells cultured in the presence or absence of the growth supporting substance "insulin like growth factor 1" (IGF-1) analogue LongR(3). Its effect on the cells ranged from enhanced growth to delay of cell death as well as cytoskeletal installation. Suggesting that under supplementation, a minimised cellular effort in installation of a large cytoskeleton occurs, possibly in favour of promoting faster cell division.
Insights
A new CHO41K microarray was developed for Chinese hamster ovary (CHO) cells, enabling detailed gene expression studies. This tool revealed that insulin-like growth factor 1 (IGF-1) supplementation enhances CHO cell growth and delays death, potentially by reducing cytoskeletal investment.
Area of Science:
- Biotechnology
- Cell Biology
- Genomics
Background:
- Chinese hamster ovary (CHO) cells are crucial for pharmaceutical protein production.
- Limited sequence information previously hindered comprehensive gene expression studies in CHO cells.
Purpose of the Study:
- To develop a high-density microarray for CHO cells to facilitate gene expression analysis.
- To investigate the impact of insulin-like growth factor 1 (IGF-1) analogue LongR(3) on CHO cell transcriptome.
Main Methods:
- Established the CHO41K microarray with 41,304 probes using existing and new sequencing data.
- Performed self-hybridization for replica determination and ran samples in triplicates for statistical power.
- Analyzed technical variance, finding no detectable intra-microarray or slide-to-slide variance.
Main Results:
- The CHO41K microarray provides reliable gene expression data with high statistical power.
- IGF-1 supplementation significantly altered CHO cell gene expression.
- Observed effects of IGF-1 included enhanced cell growth, delayed cell death, and changes in cytoskeletal installation.
Conclusions:
- The CHO41K microarray is a valuable tool for studying CHO cell biology and metabolism.
- IGF-1 supplementation may promote faster cell division by reducing cellular resources allocated to cytoskeleton formation.
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