Skip to main content

Table 3 Plasmids and strains used and constructed in this study

From: Identification and functional characterization of the CYP51 gene from the yeast Xanthophyllomyces dendrorhous that is involved in ergosterol biosynthesis

 

Relevant features or strain genotype

Source or Reference

Plasmids:

  

pBluescript SK- (pBS)

ColE1 ori; AmpR; cloning vector with blue-white selection.

Invitrogen

pMN-hph

pBS containing at the EcoRV site a cassette of 1.8 kb bearing the E. coli-Hygromycin B resistance (hph) gene under EF-1 α promoter and GPD transcription terminator of X. dendrorhous.

[56]

pBS-gCYP51

pBS bearing the genomic version of the CYP51 gene from X. dendrorhous (4,318 bp).

This work

pBS-cCYP51

pBS bearing the ORF of the CYP51 gene from X. dendrorhous (1,653 bp).

This work

pBS-cyp51/hph

pBS bearing the hygromycin B resistance cassette flanked by approximately 700 bp upstream and 600 pb downstream of the X. dendrorhous CYP51 locus.

This work

YEp-Act4

pBR322 and 2 micron ori, AmpR, LEU2 and containing the ACT promoter to regulate gene expression.

[48]

YEpNP

YEp-Act4 bearing the S. cerevisiae TDH3 terminator.

This work

YEpNP-cCYP51

YEpNP bearing the X. dendrorhous CYP51 cDNA (1,653 bp) under the regulation of the ACT promoter and the TDH3 terminator.

This work

YEpNP-gERG11

YEpNP bearing the S. cerevisiae ERG11 gene (1,593 bp) under the regulation of the ACT promoter and the TDH3 terminator.

This work

Strains:

  

E. coli:

  

DH-5α

F- φ80d lacZΔM15Δ (lacZY-argF) U169 deoR recA1 endA1 hsdR17(rk- mk+) phoA supE44 l- thi-1 gyrA96 relA1

[47]

S. cerevisiae:

  

S288c

MATα, SUC2, gal2, mal, mel, flo1, flo8-1, hap1, ho, bio1, bio6.

[64]

Meyen ex E.C Hansen YHR007C BY4743 (Sc-erg11+/−)

MATa/MATα, his3Δ1/ his3Δ1, leu2Δ0/leu2Δ0, lys2Δ0/+, met15Δ0/+, ura3Δ0/ura3Δ0, ΔERG11:KanMx. ATCC 26604.

ATCC 26604

Sc-cCYP51Xd

Diploid transformant derived from Sc-erg11 +/− containing the expression vector YEpNP-cCYP51.

This work

Sc-gERG11Sc

Diploid transformant derived from Sc-erg11 +/− containing the expression vector YEpNP-gERG11.

This work

Sc-hCYP51

Sporulation product derived from Sc-erg11 +/− containing the expression vector YEpNP- cCYP51.

This work

Sc-hERG11

Sporulation product derived from Sc-erg11 +/− containing the expression vector YEpNP-gERG11.

This work

X. dendrorhous:

  

UCD 67-385

ATCC 24230, wild-type.

ATCC 24230

CBS 6938

ATCC 96594, wild-type.

ATCC 96594

CBS-CYP51 +/−

(CBS-CYP51/cyp51 hph). Transformant derived from CBS 6938 containing an allele of the CYP51 locus replaced by a hygromycin B resistance module.

This work