Insights into the prevalence and underlying causes of clonal variation through transcriptomic analysis in Pichia pastoris

Rochelle Aw, Geraint R Barton, David J Leak

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Clonal variation, wherein a range of specific productivities of secreted proteins are observed from supposedly identical transformants, is an accepted aspect of working with Pichia pastoris. It means that a significant number of transformants need to be tested to obtain a representative sample, and in commercial protein production, companies regularly screen thousands of transformants to select for the highest secretor. Here, we have undertaken a detailed investigation of this phenomenon by characterising clones transformed with the human serum albumin gene. The titers of nine clones, each containing a single copy of the human serum albumin gene (identified by qPCR), were measured and the clones grouped into three categories, namely, high-, mid- and low-level secretors. Transcriptomic analysis, using microarrays, showed that no regulatory patterns consistently correlated with titer, suggesting that the causes of clonal variation are varied. However, a number of physiological changes appeared to underlie the differences in titer, suggesting there is more than one biochemical signature for a high-secreting strain. An anomalous low-secreting strain displaying high transcript levels that appeared to be nutritionally starved further emphasises the complicated nature of clonal variation.

LanguageEnglish
Pages5045-5058
JournalApplied Microbiology and Biotechnology
Volume101
Issue number12
Early online date22 May 2017
DOIs
StatusPublished - Jun 2017

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Pichia
Clone Cells
Serum Albumin
Microarray Analysis
Genes
Proteins

Keywords

  • Journal Article

Cite this

Insights into the prevalence and underlying causes of clonal variation through transcriptomic analysis in Pichia pastoris. / Aw, Rochelle; Barton, Geraint R; Leak, David J.

In: Applied Microbiology and Biotechnology, Vol. 101, No. 12, 06.2017, p. 5045-5058.

Research output: Contribution to journalArticle

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