A transformation system for the nonuniversal CUGSer codon usage species Candida rugosa

Shye Jye Tang; Kuang Hui Sun; Guang Huan Sun; Terng Yuan Chang; Wan Lin Wu; Guan Chiun Lee

doi:10.1016/S0167-7012(02)00182-3

A transformation system for the nonuniversal CUG^Ser codon usage species Candida rugosa

Shye Jye Tang^*, Kuang Hui Sun, Guang Huan Sun, Terng Yuan Chang, Wan Lin Wu, Guan Chiun Lee

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

12 Citations (Scopus)

Abstract

Since Candida rugosa utilizes a nonuniversal serine codon CUG rather than leucine, no vectors have been constructed to transform this organism. Moreover, it is difficult to design a new transformation system because no selection markers and promoters are available. In this study, Zeocin (400 μg/ml) was demonstrated to inhibit the growth of C. rugosa. The dominant selectable marker bleomycin-resistant determinant (ble) gene containing five CUG codons in an open-reading frame of 375 bp was synthesized by replacing its CUG codons into leucine codons (zeo-n). This marker conferred resistance to Zeocin. GAL1 promoter, transcription elongation factor 1 (TEF1) promoter from Saccharomyces cerevisiae and LIP3 promoter from C. rugosa were then used to drive zeo-n and to examine the function of promoter in C. rugosa. The resulting vectors enabled selection of Zeocin-resistant clones after transformation by LiCl method and electroporation. These results demonstrate that transformation into C. rugosa is feasible under the operation of GAL1, TEF1, and LIP3 promoters. The development of the transformation system for C. rugosa is essential to the genetic analysis of gene regulation and biochemical features of this fungal species and the expression of recombinant proteins in C. rugosa.

Original language	English
Pages (from-to)	231-238
Number of pages	8
Journal	Journal of Microbiological Methods
Volume	52
Issue number	2
DOIs	https://doi.org/10.1016/S0167-7012(02)00182-3
Publication status	Published - 2003 Feb 1
Externally published	Yes

Keywords

Bleomycin-resistant determinant (ble)
Candida rugosa
Codon usage
Dominant selection
Transformation
Zeocin

ASJC Scopus subject areas

Microbiology
Molecular Biology
Microbiology (medical)

Access to Document

10.1016/S0167-7012(02)00182-3

Cite this

@article{73ee65be66844906976c1755ff0aa890,

title = "A transformation system for the nonuniversal CUGSer codon usage species Candida rugosa",

abstract = "Since Candida rugosa utilizes a nonuniversal serine codon CUG rather than leucine, no vectors have been constructed to transform this organism. Moreover, it is difficult to design a new transformation system because no selection markers and promoters are available. In this study, Zeocin (400 μg/ml) was demonstrated to inhibit the growth of C. rugosa. The dominant selectable marker bleomycin-resistant determinant (ble) gene containing five CUG codons in an open-reading frame of 375 bp was synthesized by replacing its CUG codons into leucine codons (zeo-n). This marker conferred resistance to Zeocin. GAL1 promoter, transcription elongation factor 1 (TEF1) promoter from Saccharomyces cerevisiae and LIP3 promoter from C. rugosa were then used to drive zeo-n and to examine the function of promoter in C. rugosa. The resulting vectors enabled selection of Zeocin-resistant clones after transformation by LiCl method and electroporation. These results demonstrate that transformation into C. rugosa is feasible under the operation of GAL1, TEF1, and LIP3 promoters. The development of the transformation system for C. rugosa is essential to the genetic analysis of gene regulation and biochemical features of this fungal species and the expression of recombinant proteins in C. rugosa.",

keywords = "Bleomycin-resistant determinant (ble), Candida rugosa, Codon usage, Dominant selection, Transformation, Zeocin",

author = "Tang, {Shye Jye} and Sun, {Kuang Hui} and Sun, {Guang Huan} and Chang, {Terng Yuan} and Wu, {Wan Lin} and Lee, {Guan Chiun}",

note = "Funding Information: This work was supported by grants (NSC-89-2313-B-019-051, NSC-90-2313-B-019-028 and NSC-91-2313-B-019-040) from the National Science Council, Republic of China.",

year = "2003",

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doi = "10.1016/S0167-7012(02)00182-3",

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T1 - A transformation system for the nonuniversal CUGSer codon usage species Candida rugosa

AU - Tang, Shye Jye

AU - Sun, Kuang Hui

AU - Sun, Guang Huan

AU - Chang, Terng Yuan

AU - Wu, Wan Lin

AU - Lee, Guan Chiun

N1 - Funding Information: This work was supported by grants (NSC-89-2313-B-019-051, NSC-90-2313-B-019-028 and NSC-91-2313-B-019-040) from the National Science Council, Republic of China.

PY - 2003/2/1

Y1 - 2003/2/1

N2 - Since Candida rugosa utilizes a nonuniversal serine codon CUG rather than leucine, no vectors have been constructed to transform this organism. Moreover, it is difficult to design a new transformation system because no selection markers and promoters are available. In this study, Zeocin (400 μg/ml) was demonstrated to inhibit the growth of C. rugosa. The dominant selectable marker bleomycin-resistant determinant (ble) gene containing five CUG codons in an open-reading frame of 375 bp was synthesized by replacing its CUG codons into leucine codons (zeo-n). This marker conferred resistance to Zeocin. GAL1 promoter, transcription elongation factor 1 (TEF1) promoter from Saccharomyces cerevisiae and LIP3 promoter from C. rugosa were then used to drive zeo-n and to examine the function of promoter in C. rugosa. The resulting vectors enabled selection of Zeocin-resistant clones after transformation by LiCl method and electroporation. These results demonstrate that transformation into C. rugosa is feasible under the operation of GAL1, TEF1, and LIP3 promoters. The development of the transformation system for C. rugosa is essential to the genetic analysis of gene regulation and biochemical features of this fungal species and the expression of recombinant proteins in C. rugosa.

AB - Since Candida rugosa utilizes a nonuniversal serine codon CUG rather than leucine, no vectors have been constructed to transform this organism. Moreover, it is difficult to design a new transformation system because no selection markers and promoters are available. In this study, Zeocin (400 μg/ml) was demonstrated to inhibit the growth of C. rugosa. The dominant selectable marker bleomycin-resistant determinant (ble) gene containing five CUG codons in an open-reading frame of 375 bp was synthesized by replacing its CUG codons into leucine codons (zeo-n). This marker conferred resistance to Zeocin. GAL1 promoter, transcription elongation factor 1 (TEF1) promoter from Saccharomyces cerevisiae and LIP3 promoter from C. rugosa were then used to drive zeo-n and to examine the function of promoter in C. rugosa. The resulting vectors enabled selection of Zeocin-resistant clones after transformation by LiCl method and electroporation. These results demonstrate that transformation into C. rugosa is feasible under the operation of GAL1, TEF1, and LIP3 promoters. The development of the transformation system for C. rugosa is essential to the genetic analysis of gene regulation and biochemical features of this fungal species and the expression of recombinant proteins in C. rugosa.

KW - Bleomycin-resistant determinant (ble)

KW - Candida rugosa

KW - Codon usage

KW - Dominant selection

KW - Transformation

KW - Zeocin

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