Abstract
Preparation and regeneration of protoplasts is essential for somatic hybridization and transformation of yeasts. We present conditions that were found to be optimal for preparing and regenerating Schizosaccharomyces pombe protoplasts for cell fusion. In contrast to these conditions, genetic transformation of S. pombe requires spheroplasts that are osmotically sensitive, but still have some wall material attached to the cell. The main finding were as follows: (a) For protoplast formation with Novozym SP234, 0.9 M sorbitol was found to be the optimal osmotic milieu and β-mercaptoethanol is not necessary. (b) Embedding in soft agar yields considerably better regeneration frequencies than direct plating. (c) Cell fusion is optimal when both fusion partners are fully protoplasted, although considerable fusion occurs between spheroplasted cells as well. (d)Schizosaccharomyces pombe transformation frequencies are much higher with spheroplasts than with protoplasts. Inclusion of β-mercaptoethanol did not enhance transformation frequency.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 169-173 |
| Number of pages | 5 |
| Journal | Current Microbiology |
| Volume | 12 |
| Issue number | 3 |
| DOIs | |
| State | Published - May 1985 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Microbiology
Cite this
Preparation and regeneration of protoplasts and spheroplasts for fusion and transformation of Schizosaccharomyces pombe. / Sipiczki, Mátyás; Heyer, Wolf Dietrich; Kohli, Jürg.
In: Current Microbiology, Vol. 12, No. 3, 05.1985, p. 169-173.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Preparation and regeneration of protoplasts and spheroplasts for fusion and transformation of Schizosaccharomyces pombe
AU - Sipiczki, Mátyás
AU - Heyer, Wolf Dietrich
AU - Kohli, Jürg
PY - 1985/5
Y1 - 1985/5
N2 - Preparation and regeneration of protoplasts is essential for somatic hybridization and transformation of yeasts. We present conditions that were found to be optimal for preparing and regenerating Schizosaccharomyces pombe protoplasts for cell fusion. In contrast to these conditions, genetic transformation of S. pombe requires spheroplasts that are osmotically sensitive, but still have some wall material attached to the cell. The main finding were as follows: (a) For protoplast formation with Novozym SP234, 0.9 M sorbitol was found to be the optimal osmotic milieu and β-mercaptoethanol is not necessary. (b) Embedding in soft agar yields considerably better regeneration frequencies than direct plating. (c) Cell fusion is optimal when both fusion partners are fully protoplasted, although considerable fusion occurs between spheroplasted cells as well. (d)Schizosaccharomyces pombe transformation frequencies are much higher with spheroplasts than with protoplasts. Inclusion of β-mercaptoethanol did not enhance transformation frequency.
AB - Preparation and regeneration of protoplasts is essential for somatic hybridization and transformation of yeasts. We present conditions that were found to be optimal for preparing and regenerating Schizosaccharomyces pombe protoplasts for cell fusion. In contrast to these conditions, genetic transformation of S. pombe requires spheroplasts that are osmotically sensitive, but still have some wall material attached to the cell. The main finding were as follows: (a) For protoplast formation with Novozym SP234, 0.9 M sorbitol was found to be the optimal osmotic milieu and β-mercaptoethanol is not necessary. (b) Embedding in soft agar yields considerably better regeneration frequencies than direct plating. (c) Cell fusion is optimal when both fusion partners are fully protoplasted, although considerable fusion occurs between spheroplasted cells as well. (d)Schizosaccharomyces pombe transformation frequencies are much higher with spheroplasts than with protoplasts. Inclusion of β-mercaptoethanol did not enhance transformation frequency.
UR - http://www.scopus.com/inward/record.url?scp=0021927111&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0021927111&partnerID=8YFLogxK
U2 - 10.1007/BF01567671
DO - 10.1007/BF01567671
M3 - Article
AN - SCOPUS:0021927111
VL - 12
SP - 169
EP - 173
JO - Current Microbiology
JF - Current Microbiology
SN - 0343-8651
IS - 3
ER -