I think you would have to analyze if one of the elements of the environment in which you are working inhibits growth. Why use this medium and not another? Also I think you should analyze Pichia reaction stoichiometry and optimize...

Metab Eng. 2010 Nov;12(6):573-80. 2010 Aug 3.

Engineering of biotin-prototrophy in Pichia pastoris for robust production processes.

Gasser B1, Dragosits M, Mattanovich D.

Abstract

Biotin plays an essential role as cofactor for biotin-dependent carboxylases involved in essential metabolic pathways. The cultivation of Pichia pastoris, a methylotrophic yeast that is successfully used as host for the production of recombinant proteins, requires addition of high dosage of biotin. As biotin is the only non-salt media component used during P. pastoris fermentation (apart from the carbon source), nonconformities during protein production processes are usually attributed to poor quality of the added biotin. In order to avoid dismissed production runs due to biotin quality issues, we engineered the biotin-requiring yeast P. pastoris to become a biotin-prototrophic yeast. Integration of four genes involved in the biotin biosynthesis from brewing yeast into the P. pastoris genome rendered P. pastoris biotin-prototrophic. The engineered strain has successfully been used as production host for both intracellular and secreted heterologous proteins in fed-batch processes, employing mineral media without vitamins. Another field of application for these truly prototrophic hosts is the production of biochemicals and small metabolites, where defined mineral media leads to easier purification procedures.

Hi there,

To my point Pichia always needs biotin for groowing

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10/5/17

We grow Pichia pastoris in our 5-L fermentors from time to time. We use Invitrogen's "Expression Media, Fermentation Basal Salts". This is a special formulation for Pichia and it works well. Invitrogen doesn't list the composition, so I don't know if it contains biotin, but you might try it. We have had good luck with it.

I hope this information helps you.

Bill Colonna  Center for Crops Utilization Research, Iowa State University, Ames, IA,  USA    [email protected].

Thank you.

Do you use pH 5?

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10/5/17

Yes. The initial pH of the medium is

Hi, Which strain are you using ?. X-33, GS115 or something else.....?

Check the clone history, if it is growing in complex medium and not in minimal medium, there must be some deletion of gene in genome level which could produce some essential amino acids such as histidine...

Adjusting to a higher pH solved the issue

alternately you may also add a phosphate buffered in minimal medium will work

To answer the initial question of Biotin requirement is needed for most of the strains