EST sequencing for gene discovery in Chinese hamster ovary cells

Chinese hamster ovary (CHO) cells are one of the most important cell lines in biological research, and are the most widely used host for industrial production of recombinant therapeutic proteins. Despite their extensive applications, little sequence information is available for molecular based resea...

Full description

Saved in:
Bibliographic Details
Published in:Biotechnology and bioengineering 2005-09, Vol.91 (5), p.592-606
Main Authors: Wlaschin, Katie Fraass, Nissom, Peter Morin, Gatti, Marcela de Leon, Ong, Peh Fern, Arleen, Sanny, Tan, Kher Shing, Rink, Anette, Cham, Breana, Wong, Kathy, Yap, Miranda, Hu, Wei-Shou
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Base Sequence
Biological and medical sciences
Biotechnology
Calmodulin - chemistry
cDNA microarray
Chinese hamster ovary cells (CHO cells)
CHO Cells
Cricetinae
Cricetulus
Electron Transport Complex IV - chemistry
EST sequencing
Expressed Sequence Tags
Fundamental and applied biological sciences. Psychology
Gene Library
Gene therapy
Genome
Isoenzymes - chemistry
L-Lactate Dehydrogenase - chemistry
Mitochondria - genetics
Mitochondrial DNA
mitochondrial genome
Molecular Sequence Data
Oligonucleotide Array Sequence Analysis
Proteins
Ribosomal Proteins - chemistry
RNA, Messenger - metabolism
Sequence Analysis, DNA
Sequence Homology, Amino Acid
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Chinese hamster ovary (CHO) cells are one of the most important cell lines in biological research, and are the most widely used host for industrial production of recombinant therapeutic proteins. Despite their extensive applications, little sequence information is available for molecular based research. To facilitate gene discovery and genetic engineering, two cDNA libraries were constructed from three CHO cell lines grown under various conditions. The average insert size for both libraries is approximately 800–850 bp, and each library has comparable redundancy levels of 36%–38% for the sequences isolated. Random sequencing of 4,608 ESTs yielded 2,602 unique assemblies, 76% of which were annotated as orthologs of sequences in the GenBank database. A high abundance of mitochondrial genome transcripts facilitated the assembly of the complete mitochondrial genome by PCR walking. Comparative analysis of sequences from both mitochondrial and nuclear genomes with orthologous genes from other species shows that CHO sequences are generally most similar to mouse; however, examples with highest similarity to rat or human are common. A cDNA microarray, including all 4,608 ESTs, was constructed. The microarray results reveal a high level of consistency between transcript abundance in the libraries and fluorescence intensities. Inclusion of redundant clones in the microarray, additionally, allows small changes in abundant mRNAs to be discerned with a high degree of confidence. The information and tools generated provide access to genomic technology for this important cell line. © 2005 Wiley Periodicals, Inc.
ISSN:0006-3592
1097-0290
DOI:10.1002/bit.20511