Structured Review

Thermo Fisher pyes2
Copper Assimilation Function of Chlamydomonas CTR1 and CTR2. A Δ ctr1 mutant of S. cerevisiae was transformed with the yeast expression vector <t>pYES2</t> as negative control (1), with pYES2 carrying the S. cerevisiae CTR1 gene as positive control (2), and with pYES2 carrying cDNAs for C. reinhardtii CTR1 (3), CTR2 (4), COPT1 (5), and CTR3 (6) as the test constructs. Transformants were streaked to media containing ethanol (3% v/v) as carbon source (YPE) to test for respiratory competence and to YPE containing 100 μM CuSO 4 as a control. Media were supplemented with galactose (0.02%; see Methods). Plates were incubated at 30°C and photographed at 9 and 7 d, respectively.
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1) Product Images from "Two Chlamydomonas CTR Copper Transporters with a Novel Cys-Met Motif Are Localized to the Plasma Membrane and Function in Copper Assimilation"

Article Title: Two Chlamydomonas CTR Copper Transporters with a Novel Cys-Met Motif Are Localized to the Plasma Membrane and Function in Copper Assimilation

Journal: The Plant Cell

doi: 10.1105/tpc.108.064907

Copper Assimilation Function of Chlamydomonas CTR1 and CTR2. A Δ ctr1 mutant of S. cerevisiae was transformed with the yeast expression vector pYES2 as negative control (1), with pYES2 carrying the S. cerevisiae CTR1 gene as positive control (2), and with pYES2 carrying cDNAs for C. reinhardtii CTR1 (3), CTR2 (4), COPT1 (5), and CTR3 (6) as the test constructs. Transformants were streaked to media containing ethanol (3% v/v) as carbon source (YPE) to test for respiratory competence and to YPE containing 100 μM CuSO 4 as a control. Media were supplemented with galactose (0.02%; see Methods). Plates were incubated at 30°C and photographed at 9 and 7 d, respectively.
Figure Legend Snippet: Copper Assimilation Function of Chlamydomonas CTR1 and CTR2. A Δ ctr1 mutant of S. cerevisiae was transformed with the yeast expression vector pYES2 as negative control (1), with pYES2 carrying the S. cerevisiae CTR1 gene as positive control (2), and with pYES2 carrying cDNAs for C. reinhardtii CTR1 (3), CTR2 (4), COPT1 (5), and CTR3 (6) as the test constructs. Transformants were streaked to media containing ethanol (3% v/v) as carbon source (YPE) to test for respiratory competence and to YPE containing 100 μM CuSO 4 as a control. Media were supplemented with galactose (0.02%; see Methods). Plates were incubated at 30°C and photographed at 9 and 7 d, respectively.

Techniques Used: Mutagenesis, Transformation Assay, Expressing, Plasmid Preparation, Negative Control, Positive Control, Construct, Incubation

2) Product Images from "Heterologous expression of an α-amylase inhibitor from common bean (Phaseolus vulgaris) in Kluyveromyces lactis and Saccharomyces cerevisiae"

Article Title: Heterologous expression of an α-amylase inhibitor from common bean (Phaseolus vulgaris) in Kluyveromyces lactis and Saccharomyces cerevisiae

Journal: Microbial Cell Factories

doi: 10.1186/s12934-017-0719-4

Schematic representation of the gene constructs used for αAI production. Diagrams of a the native αAI gene from cv. Pinto comprising signal peptide (SP), α- and β- subunits and the propeptide sequences; b the synthetic αAI - OPT gene, optimised for expression in two yeasts, including a Kozak consensus sequence and flanked by Hind III and Not I restriction sites. Diagrams of constructs c pKLAC2-αAI-OPT and d pYES2-αAI-OPT. P Lac4-PBI and P Gal1 correspond to promoters Lac4-PBI and Gal1 in pKLAC2 and pYES2, respectively. TT Lac4-PBI and TT CYC correspond to 3′ terminators Lac4-PBI and CYC in pKLAC2 and pYES2, respectively
Figure Legend Snippet: Schematic representation of the gene constructs used for αAI production. Diagrams of a the native αAI gene from cv. Pinto comprising signal peptide (SP), α- and β- subunits and the propeptide sequences; b the synthetic αAI - OPT gene, optimised for expression in two yeasts, including a Kozak consensus sequence and flanked by Hind III and Not I restriction sites. Diagrams of constructs c pKLAC2-αAI-OPT and d pYES2-αAI-OPT. P Lac4-PBI and P Gal1 correspond to promoters Lac4-PBI and Gal1 in pKLAC2 and pYES2, respectively. TT Lac4-PBI and TT CYC correspond to 3′ terminators Lac4-PBI and CYC in pKLAC2 and pYES2, respectively

Techniques Used: Construct, Expressing, Sequencing

Western blot and α-amylase inhibitory activity of proteins from selected colonies of S. cerevisiae YPH499 with αAI - OPT . a Western blot of proteins (25 µg) from culture supernatants and lysed-cells of colonies 8, 27 and 92 transformed with αAI - OPT ; the parent strain YPH499 transformed with empty plasmid pYES2 was used as negative control (−C) and pure Pinto-αAI (200 ng) was used as positive control (+C). Molecular markers are shown in kilodaltons at left . b Inhibitory activity against porcine pancreatic α-amylase of lysed-cell extracts (100 µg of protein) of S. cerevisiae YPH499/pYES2-αAI-OPT colonies 8, 27 and 92. Parent strain YPH499 transformed with empty plasmid pYES2 was used as a negative control (−C) and pure Pinto-αAI (200 ng) was used as a positive control (+C). Results are expressed as inhibition units (IU). Data from triplicate experiments were expressed as the average ± standard error. *** P
Figure Legend Snippet: Western blot and α-amylase inhibitory activity of proteins from selected colonies of S. cerevisiae YPH499 with αAI - OPT . a Western blot of proteins (25 µg) from culture supernatants and lysed-cells of colonies 8, 27 and 92 transformed with αAI - OPT ; the parent strain YPH499 transformed with empty plasmid pYES2 was used as negative control (−C) and pure Pinto-αAI (200 ng) was used as positive control (+C). Molecular markers are shown in kilodaltons at left . b Inhibitory activity against porcine pancreatic α-amylase of lysed-cell extracts (100 µg of protein) of S. cerevisiae YPH499/pYES2-αAI-OPT colonies 8, 27 and 92. Parent strain YPH499 transformed with empty plasmid pYES2 was used as a negative control (−C) and pure Pinto-αAI (200 ng) was used as a positive control (+C). Results are expressed as inhibition units (IU). Data from triplicate experiments were expressed as the average ± standard error. *** P

Techniques Used: Western Blot, Activity Assay, Transformation Assay, Plasmid Preparation, Negative Control, Positive Control, Inhibition

3) Product Images from "The Thellungiella salsuginea Tonoplast Aquaporin TsTIP1;2 Functions in Protection Against Multiple Abiotic Stresses"

Article Title: The Thellungiella salsuginea Tonoplast Aquaporin TsTIP1;2 Functions in Protection Against Multiple Abiotic Stresses

Journal: Plant and Cell Physiology

doi: 10.1093/pcp/pct166

H 2 O 2 permeability of yeast cells expressing TsTIP1;2 . (A) Saccharomyces cerevisiae aqy-null strain cells transformed with the empty vector pYES2 alone or pYES2-TsTIP1;2 were spotted in 10-fold dilutions on medium without or with 1, 1.5 or 2 mM H 2 O 2 , respectively.
Figure Legend Snippet: H 2 O 2 permeability of yeast cells expressing TsTIP1;2 . (A) Saccharomyces cerevisiae aqy-null strain cells transformed with the empty vector pYES2 alone or pYES2-TsTIP1;2 were spotted in 10-fold dilutions on medium without or with 1, 1.5 or 2 mM H 2 O 2 , respectively.

Techniques Used: Permeability, Expressing, Transformation Assay, Plasmid Preparation

4) Product Images from "Pxl1p, a Paxillin-like Protein in Saccharomyces cerevisiae, May Coordinate Cdc42p and Rho1p Functions during Polarized Growth"

Article Title: Pxl1p, a Paxillin-like Protein in Saccharomyces cerevisiae, May Coordinate Cdc42p and Rho1p Functions during Polarized Growth

Journal: Molecular Biology of the Cell

doi: 10.1091/mbc.E04-01-0079

Antagonistic relationship between Cdc42p and Rho1p signaling pathways. (A) Overexpression of Cdc42p inhibits the growth of rho1- Ts mutants. Yeast strains NY2284 (WT), NY2285 ( rho1-2 ) and NY2286 ( rho1-3 ) carrying plasmid YEp352 (Vec) or YEp352-GAL-CDC42 were streaked onto SC-Ura plates containing dextrose (Dextrose) and SC-Ura plates containing 2% galactose and 1% raffinose (Galactose), and incubated at 30°C for 3 days. (B) Overexpression of Rho1p inhibits the growth of cdc42-201 , but not cdc42 G60D mutant. Yeast strains YEF473A (WT), YEF2258 ( cdc42-201 ), and JPC241 ( cdc42 G60D ) carrying plasmid pYES2 (Vec) or pYES2-RHO1 (GAL-RHO1) were streaked onto SC-Ura plates containing 2% galactose and 1% raffinose (Galactose), and incubated at 24 or 30°C for 3 days. Control SC-Ura plate containing dextrose (Dextrose) was incubated at 24°C for 3 days.
Figure Legend Snippet: Antagonistic relationship between Cdc42p and Rho1p signaling pathways. (A) Overexpression of Cdc42p inhibits the growth of rho1- Ts mutants. Yeast strains NY2284 (WT), NY2285 ( rho1-2 ) and NY2286 ( rho1-3 ) carrying plasmid YEp352 (Vec) or YEp352-GAL-CDC42 were streaked onto SC-Ura plates containing dextrose (Dextrose) and SC-Ura plates containing 2% galactose and 1% raffinose (Galactose), and incubated at 30°C for 3 days. (B) Overexpression of Rho1p inhibits the growth of cdc42-201 , but not cdc42 G60D mutant. Yeast strains YEF473A (WT), YEF2258 ( cdc42-201 ), and JPC241 ( cdc42 G60D ) carrying plasmid pYES2 (Vec) or pYES2-RHO1 (GAL-RHO1) were streaked onto SC-Ura plates containing 2% galactose and 1% raffinose (Galactose), and incubated at 24 or 30°C for 3 days. Control SC-Ura plate containing dextrose (Dextrose) was incubated at 24°C for 3 days.

Techniques Used: Over Expression, Plasmid Preparation, Incubation, Mutagenesis

5) Product Images from "The Proteasomal Substrate Stm1 Participates in Apoptosis-like Cell Death in Yeast"

Article Title: The Proteasomal Substrate Stm1 Participates in Apoptosis-like Cell Death in Yeast

Journal: Molecular Biology of the Cell

doi:

Several HEL genes cause growth arrest and cell death when overexpressed in proteasomal mutants. Wild-type and pre1-1 pre4-1 cells carrying pYES2-encoded, P GAL1 -driven cDNAs of HEL genes (as indicated) were cultivated overnight in SC ura − . Cells were harvested, and washed in water. (A) To test for growth arrest, 10-fold serial dilutions were spotted on SCgal ura − plates. Plates were incubated for 3 d at 30°C. (B) After 8 h of induction in SCgal ura − liquid medium, cell death rate was scored as 100% − (plating efficiency on YPD medium). Frequency of cells with TUNEL positive nuclei was determined by microscopic inspection. Results were averaged from two experiments.
Figure Legend Snippet: Several HEL genes cause growth arrest and cell death when overexpressed in proteasomal mutants. Wild-type and pre1-1 pre4-1 cells carrying pYES2-encoded, P GAL1 -driven cDNAs of HEL genes (as indicated) were cultivated overnight in SC ura − . Cells were harvested, and washed in water. (A) To test for growth arrest, 10-fold serial dilutions were spotted on SCgal ura − plates. Plates were incubated for 3 d at 30°C. (B) After 8 h of induction in SCgal ura − liquid medium, cell death rate was scored as 100% − (plating efficiency on YPD medium). Frequency of cells with TUNEL positive nuclei was determined by microscopic inspection. Results were averaged from two experiments.

Techniques Used: Incubation, TUNEL Assay

6) Product Images from "Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard"

Article Title: Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard

Journal: Scientific Reports

doi: 10.1038/s41598-017-08317-y

Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.
Figure Legend Snippet: Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.

Techniques Used: Expressing, Gas Chromatography, Construct, Sequencing, Plasmid Preparation

7) Product Images from "Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption"

Article Title: Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption

Journal: International Journal for Parasitology: Drugs and Drug Resistance

doi: 10.1016/j.ijpddr.2018.06.003

Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.
Figure Legend Snippet: Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.

Techniques Used: Functional Assay, Mutagenesis, Transformation Assay, Expressing, Plasmid Preparation, Positive Control

8) Product Images from "Genome-wide analysis of heat shock proteins in C4 model, foxtail millet identifies potential candidates for crop improvement under abiotic stress"

Article Title: Genome-wide analysis of heat shock proteins in C4 model, foxtail millet identifies potential candidates for crop improvement under abiotic stress

Journal: Scientific Reports

doi: 10.1038/srep32641

Spot assay of yeast (W303) cells on SD/-ura basal medium. Growth of control pYES2 and Sishsp27 -pYES2 transformed yeast cells under different stress conditions.
Figure Legend Snippet: Spot assay of yeast (W303) cells on SD/-ura basal medium. Growth of control pYES2 and Sishsp27 -pYES2 transformed yeast cells under different stress conditions.

Techniques Used: Spot Test, Transformation Assay

9) Product Images from "The Pmt2p-Mediated Protein O-Mannosylation Is Required for Morphogenesis, Adhesive Properties, Cell Wall Integrity and Full Virulence of Magnaporthe oryzae"

Article Title: The Pmt2p-Mediated Protein O-Mannosylation Is Required for Morphogenesis, Adhesive Properties, Cell Wall Integrity and Full Virulence of Magnaporthe oryzae

Journal: Frontiers in Microbiology

doi: 10.3389/fmicb.2016.00630

M. oryzae MoPmt2 encodes a functional homolog of S. cerevisiae Pmt2. (A) Prediction of transmembrane structure in MoPmt2. The position of each transmembrane domain was generated by the TMHMM Server v. 2.0 online program, and the MoPmt2 possess the typical hydropathy profiles of O -mannosyltransferases. (B) Conservation of the Pmt sequence motifs in MoPmt2. Physical map of MoPmt2 revealed two conserved motifs (Pmt and MIR) of the PMT family at the N-terminal transmembrane region and central hydrophilic region. (C) Functional complementation of S. cerevisiae Pmt2 mutant. Serial 10-fold dilutions of cells from wild-type strain BY4741 transformed with pYES2 and the Pmt2 strain transformed with either pYES2 or pYES2- M oPmt2 are dotted onto SC-Ura plates with Glu or Gal supplemented with 12.5 μg/mL CFW as indicated, and incubated at 30°C for 72 h.
Figure Legend Snippet: M. oryzae MoPmt2 encodes a functional homolog of S. cerevisiae Pmt2. (A) Prediction of transmembrane structure in MoPmt2. The position of each transmembrane domain was generated by the TMHMM Server v. 2.0 online program, and the MoPmt2 possess the typical hydropathy profiles of O -mannosyltransferases. (B) Conservation of the Pmt sequence motifs in MoPmt2. Physical map of MoPmt2 revealed two conserved motifs (Pmt and MIR) of the PMT family at the N-terminal transmembrane region and central hydrophilic region. (C) Functional complementation of S. cerevisiae Pmt2 mutant. Serial 10-fold dilutions of cells from wild-type strain BY4741 transformed with pYES2 and the Pmt2 strain transformed with either pYES2 or pYES2- M oPmt2 are dotted onto SC-Ura plates with Glu or Gal supplemented with 12.5 μg/mL CFW as indicated, and incubated at 30°C for 72 h.

Techniques Used: Functional Assay, Generated, Sequencing, Mutagenesis, Transformation Assay, Incubation

10) Product Images from "Saccharomyces cerevisiae ?1278b Has Novel Genes of the N-Acetyltransferase Gene Superfamily Required for l-Proline Analogue Resistance"

Article Title: Saccharomyces cerevisiae ?1278b Has Novel Genes of the N-Acetyltransferase Gene Superfamily Required for l-Proline Analogue Resistance

Journal: Journal of Bacteriology

doi:

Growth phenotype of S. cerevisiae strains on minimal medium containing AZC. Approximately 10 6 cells of each strain and serial dilutions of 10 −1 to 10 −3 (from left to right) were spotted onto SD plates with appropriate amino acids in the absence (−AZC) and presence (+AZC) of AZC. Plates were incubated at 30°C for 3 days. (A) Function of the MPR1 and MPR2 genes in Σ1278b background strains. The MPR1 and MPR2 disruptants derived from strain FH506 are represented by FH506D1 and FH506D2, respectively. The MPR1 MPR2 double disruptant is represented by FH506D12. (B) Function of the MPR1 gene in the other S. cerevisiae strains. Plasmids pMH1 and pMH3 were constructed from vectors pYES2 and pRS406, respectively.
Figure Legend Snippet: Growth phenotype of S. cerevisiae strains on minimal medium containing AZC. Approximately 10 6 cells of each strain and serial dilutions of 10 −1 to 10 −3 (from left to right) were spotted onto SD plates with appropriate amino acids in the absence (−AZC) and presence (+AZC) of AZC. Plates were incubated at 30°C for 3 days. (A) Function of the MPR1 and MPR2 genes in Σ1278b background strains. The MPR1 and MPR2 disruptants derived from strain FH506 are represented by FH506D1 and FH506D2, respectively. The MPR1 MPR2 double disruptant is represented by FH506D12. (B) Function of the MPR1 gene in the other S. cerevisiae strains. Plasmids pMH1 and pMH3 were constructed from vectors pYES2 and pRS406, respectively.

Techniques Used: Incubation, Derivative Assay, Construct

Restriction map of the cloned DNA fragment and deletion analysis to identify the region required for AZC resistance. The predicted size and transcriptional orientation of each deduced open reading frame (ORF) is shown by an arrow. Restriction enzymes: Ba, Bam HI; Bg, Bgl II; Ec, Eco RI; Hi, Hin dIII; Ml, Mlu I; Sa, Sac I; Su, Sau 3AI; Xh, Xho I. (A) Restriction map of the cloned DNA fragment in pMH1 and pMH2. Two plasmids had the overlapping 5.4-kb Sau 3AI insert (open box). The region in each plasmid matching the sequence on chromosome XIV (pMH1) or X (pMH2) of S. cerevisiae S288C is indicated by a shaded box. The hatched box in pMH2 represents the unknown, partially sequenced 4.6-kb fragment. (B) Analysis of MPR1 deletion mutants. Each DNA fragment was subcloned into pYES2, and the resultant plasmids were introduced into strain CKY2. AZC resistance of the Ura + transformants was examined on SD agar plates containing AZC (0.3 mg/ml) after incubation at 30°C for 3 days. +, growth; −, no growth.
Figure Legend Snippet: Restriction map of the cloned DNA fragment and deletion analysis to identify the region required for AZC resistance. The predicted size and transcriptional orientation of each deduced open reading frame (ORF) is shown by an arrow. Restriction enzymes: Ba, Bam HI; Bg, Bgl II; Ec, Eco RI; Hi, Hin dIII; Ml, Mlu I; Sa, Sac I; Su, Sau 3AI; Xh, Xho I. (A) Restriction map of the cloned DNA fragment in pMH1 and pMH2. Two plasmids had the overlapping 5.4-kb Sau 3AI insert (open box). The region in each plasmid matching the sequence on chromosome XIV (pMH1) or X (pMH2) of S. cerevisiae S288C is indicated by a shaded box. The hatched box in pMH2 represents the unknown, partially sequenced 4.6-kb fragment. (B) Analysis of MPR1 deletion mutants. Each DNA fragment was subcloned into pYES2, and the resultant plasmids were introduced into strain CKY2. AZC resistance of the Ura + transformants was examined on SD agar plates containing AZC (0.3 mg/ml) after incubation at 30°C for 3 days. +, growth; −, no growth.

Techniques Used: Clone Assay, Plasmid Preparation, Sequencing, Incubation

11) Product Images from "Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption"

Article Title: Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption

Journal: International Journal for Parasitology: Drugs and Drug Resistance

doi: 10.1016/j.ijpddr.2018.06.003

Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.
Figure Legend Snippet: Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.

Techniques Used: Functional Assay, Mutagenesis, Transformation Assay, Expressing, Plasmid Preparation, Positive Control

12) Product Images from "Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard"

Article Title: Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard

Journal: Scientific Reports

doi: 10.1038/s41598-017-08317-y

Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.
Figure Legend Snippet: Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.

Techniques Used: Expressing, Gas Chromatography, Construct, Sequencing, Plasmid Preparation

13) Product Images from "Improved Cd, Zn and Mn tolerance and reduced Cd accumulation in grains with wheat-based cell number regulator TaCNR2"

Article Title: Improved Cd, Zn and Mn tolerance and reduced Cd accumulation in grains with wheat-based cell number regulator TaCNR2

Journal: Scientific Reports

doi: 10.1038/s41598-018-37352-6

TaCNR2 -transgenic yeast grown under heavy metal treatments. The pYES2 and pYES2- TaCNR2 were transformed into yeast strain (BY4741). Gradient dilution was spotted onto the solid media (yeast extract; peptone; galactose) with 50 μM CdSO 4 , 5 mM ZnSO 4 and 5 mM MnSO 4 . The YPD medium (yeast extract; peptone; glucose) was the control. Growth was maintained at 30 °C for 3–7 days.
Figure Legend Snippet: TaCNR2 -transgenic yeast grown under heavy metal treatments. The pYES2 and pYES2- TaCNR2 were transformed into yeast strain (BY4741). Gradient dilution was spotted onto the solid media (yeast extract; peptone; galactose) with 50 μM CdSO 4 , 5 mM ZnSO 4 and 5 mM MnSO 4 . The YPD medium (yeast extract; peptone; glucose) was the control. Growth was maintained at 30 °C for 3–7 days.

Techniques Used: Transgenic Assay, Transformation Assay

14) Product Images from "Overexpression of MfPIP2-7 from Medicago falcata promotes cold tolerance and growth under NO3− deficiency in transgenic tobacco plants"

Article Title: Overexpression of MfPIP2-7 from Medicago falcata promotes cold tolerance and growth under NO3− deficiency in transgenic tobacco plants

Journal: BMC Plant Biology

doi: 10.1186/s12870-016-0814-4

Yeast growth and survival test on medium containing H 2 O 2 . After a series of dilution of the yeast cells transformed with either an empty pYES2 as control or derivate of pYES2 carrying MfPIP2-7 (pYES2-PIP2-7) at an A 600nm of 0.6, 10 μl was spotted on medium containing various concentrations of H 2 O 2 as indicated. Growth was recorded after 4 days at 30 °C
Figure Legend Snippet: Yeast growth and survival test on medium containing H 2 O 2 . After a series of dilution of the yeast cells transformed with either an empty pYES2 as control or derivate of pYES2 carrying MfPIP2-7 (pYES2-PIP2-7) at an A 600nm of 0.6, 10 μl was spotted on medium containing various concentrations of H 2 O 2 as indicated. Growth was recorded after 4 days at 30 °C

Techniques Used: Transformation Assay

15) Product Images from "Plasma membrane-localized transporter for aluminum in rice"

Article Title: Plasma membrane-localized transporter for aluminum in rice

Journal: Proceedings of the National Academy of Sciences of the United States of America

doi: 10.1073/pnas.1004949107

Transport activity of Nrat1 for aluminum in yeast. ( A ) Effect of Nrat1 expression on Al tolerance. Yeast cells (BY4741) carrying empty vector pYES2, Nrat1 , or AtNramp4 were spotted on LPM-ura medium (pH 4.2) buffered with 5 mM succinic acid with or without AlCl 3 at different dilutions. The plates were incubated at 30 °C for 2 to 3 d. ( B ) Transport activity of Nrat1 for trivalent Al ion. Yeast cells carrying empty vector pYES2, Nrat1 , or AtNramp4 were exposed to 10, 30, 50, or 100 μM AlCl 3 at pH 4.2 for 2 h. ( C ) Time-dependent transport of Nrat1 for Al. Yeast cells carrying empty vector pYES2 and Nrat1 were exposed to 50 μM AlCl 3 at pH 4.2 for different times. Net uptake was the difference between Al uptake from yeast carrying Nrat1 and empty vector. ( D ) Transport activity of Nrat1 for different Al forms. Yeast cells transformed with Nrat1 were exposed for 2 h to a solution (pH 4.2) containing 50 μM AlCl 3 , or an Al–citrate complex prepared by mixing 50 μM AlCl 3 with 500 μM citrate. After uptake, the yeast cells were washed and digested with HCl. The aluminum concentration in the digest solution was determined by atomic absorption spectrophotometer. Data in B , C , and D are means ± SD of three biological replicates.
Figure Legend Snippet: Transport activity of Nrat1 for aluminum in yeast. ( A ) Effect of Nrat1 expression on Al tolerance. Yeast cells (BY4741) carrying empty vector pYES2, Nrat1 , or AtNramp4 were spotted on LPM-ura medium (pH 4.2) buffered with 5 mM succinic acid with or without AlCl 3 at different dilutions. The plates were incubated at 30 °C for 2 to 3 d. ( B ) Transport activity of Nrat1 for trivalent Al ion. Yeast cells carrying empty vector pYES2, Nrat1 , or AtNramp4 were exposed to 10, 30, 50, or 100 μM AlCl 3 at pH 4.2 for 2 h. ( C ) Time-dependent transport of Nrat1 for Al. Yeast cells carrying empty vector pYES2 and Nrat1 were exposed to 50 μM AlCl 3 at pH 4.2 for different times. Net uptake was the difference between Al uptake from yeast carrying Nrat1 and empty vector. ( D ) Transport activity of Nrat1 for different Al forms. Yeast cells transformed with Nrat1 were exposed for 2 h to a solution (pH 4.2) containing 50 μM AlCl 3 , or an Al–citrate complex prepared by mixing 50 μM AlCl 3 with 500 μM citrate. After uptake, the yeast cells were washed and digested with HCl. The aluminum concentration in the digest solution was determined by atomic absorption spectrophotometer. Data in B , C , and D are means ± SD of three biological replicates.

Techniques Used: Activity Assay, Expressing, Plasmid Preparation, Incubation, Transformation Assay, Concentration Assay, Spectrophotometry

16) Product Images from "Metal transport protein 8 in Camellia sinensis confers superior manganese tolerance when expressed in yeast and Arabidopsis thaliana"

Article Title: Metal transport protein 8 in Camellia sinensis confers superior manganese tolerance when expressed in yeast and Arabidopsis thaliana

Journal: Scientific Reports

doi: 10.1038/srep39915

Effect of CsMTP8 expression on Mn tolerance and Mn accumulation in Saccharomyces cerevisiae . ( A ) The yeast mutants INVSC1 carrying the pYES2 empty vector or pYES2-CsMTP8 were used. Yeast cells with optical densities of 2.0 at 600 nm (OD 600 ) and samples taken after six gradient 1:10 dilutions were spotted onto synthetic complete medium containing 10 mM, and 15 mM MnSO 4 . They were left to culture for 48 h at 30 °C. EV indicates yeast cells transformed with the empty vector pYES2. OX1 and OX2 are two clones showing CsMTP8 overexpression. ( B ) Mn accumulation in wild-type strain transformed with empty vector pYES2 or pYES2-CsMTP8. Following cultured in liquid SC-U/Gal medium supplemented with 5 mM MnSO 4 at an initial OD 600 = 0.1 for 24 h. Asterisks indicate a significant difference at the P = 0.05 level compared to EV.
Figure Legend Snippet: Effect of CsMTP8 expression on Mn tolerance and Mn accumulation in Saccharomyces cerevisiae . ( A ) The yeast mutants INVSC1 carrying the pYES2 empty vector or pYES2-CsMTP8 were used. Yeast cells with optical densities of 2.0 at 600 nm (OD 600 ) and samples taken after six gradient 1:10 dilutions were spotted onto synthetic complete medium containing 10 mM, and 15 mM MnSO 4 . They were left to culture for 48 h at 30 °C. EV indicates yeast cells transformed with the empty vector pYES2. OX1 and OX2 are two clones showing CsMTP8 overexpression. ( B ) Mn accumulation in wild-type strain transformed with empty vector pYES2 or pYES2-CsMTP8. Following cultured in liquid SC-U/Gal medium supplemented with 5 mM MnSO 4 at an initial OD 600 = 0.1 for 24 h. Asterisks indicate a significant difference at the P = 0.05 level compared to EV.

Techniques Used: Expressing, Plasmid Preparation, Transformation Assay, Clone Assay, Over Expression, Cell Culture

17) Product Images from "The Evolution and Biocatalysis of FAD2 Indicate Its Correlation to the Content of Seed Oil in Plants"

Article Title: The Evolution and Biocatalysis of FAD2 Indicate Its Correlation to the Content of Seed Oil in Plants

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms20040849

The conversion of oleic acid into linoleic acid in Saccharomyces cerevisiae cells overexpressing the seed-specific or high-expressed FAD2 genes in soybean ( GmaFAD2-1 and GmaFAD2-2 ), perilla ( PfrFAD2-1 and PfrFAD2-2 ), rice ( OsaFAD2-1 ), and spruce ( PabFAD2-1 ). The control represented the conversion ratio in yeast with the empty vector pYES2. The conversion ratios were calculated based on the results of Gas Chromatography (GC) in Figure S3 of File S3 . The p -values are also presented in Figure S3 in File S3 .
Figure Legend Snippet: The conversion of oleic acid into linoleic acid in Saccharomyces cerevisiae cells overexpressing the seed-specific or high-expressed FAD2 genes in soybean ( GmaFAD2-1 and GmaFAD2-2 ), perilla ( PfrFAD2-1 and PfrFAD2-2 ), rice ( OsaFAD2-1 ), and spruce ( PabFAD2-1 ). The control represented the conversion ratio in yeast with the empty vector pYES2. The conversion ratios were calculated based on the results of Gas Chromatography (GC) in Figure S3 of File S3 . The p -values are also presented in Figure S3 in File S3 .

Techniques Used: Plasmid Preparation, Gas Chromatography

18) Product Images from "Dexamethasone Partially Rescues Ataxia Telangiectasia-mutated (ATM) Deficiency in Ataxia Telangiectasia by Promoting a Shortened Protein Variant Retaining Kinase Activity *"

Article Title: Dexamethasone Partially Rescues Ataxia Telangiectasia-mutated (ATM) Deficiency in Ataxia Telangiectasia by Promoting a Shortened Protein Variant Retaining Kinase Activity *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M112.344473

Functionality of miniATM. A , yeast complementation assay was performed with tel1 Δ (KSC1368) and mec1 -81 tel1 Δ (KSC1402) yeast mutants transfected with pYES2-ATMdexa1 or empty pYES2 constructs. Recombinants were grown on solid selective medium or liquid selective medium supplemented with the DSB inducer phleomycin ( B ) to assess the ability of ATMdexa1 to complement TEL1 and TEL1-MEC1 deficiencies. O.D. , optical density. C , miniATM kinase assay. Recombinant GST-miniATM was tested for in vitro phosphorylation of PHAS-I substrate in the presence of [γ- 32 P]ATP. Two kinase concentrations were tested, with or without wortmannin inhibition.
Figure Legend Snippet: Functionality of miniATM. A , yeast complementation assay was performed with tel1 Δ (KSC1368) and mec1 -81 tel1 Δ (KSC1402) yeast mutants transfected with pYES2-ATMdexa1 or empty pYES2 constructs. Recombinants were grown on solid selective medium or liquid selective medium supplemented with the DSB inducer phleomycin ( B ) to assess the ability of ATMdexa1 to complement TEL1 and TEL1-MEC1 deficiencies. O.D. , optical density. C , miniATM kinase assay. Recombinant GST-miniATM was tested for in vitro phosphorylation of PHAS-I substrate in the presence of [γ- 32 P]ATP. Two kinase concentrations were tested, with or without wortmannin inhibition.

Techniques Used: Transfection, Construct, Kinase Assay, Recombinant, In Vitro, Inhibition

19) Product Images from "Development of Bacteriocinogenic Strains of Saccharomyces cerevisiae Heterologously Expressing and Secreting the Leaderless Enterocin L50 Peptides L50A and L50B from Enterococcus faecium L50 ▿"

Article Title: Development of Bacteriocinogenic Strains of Saccharomyces cerevisiae Heterologously Expressing and Secreting the Leaderless Enterocin L50 Peptides L50A and L50B from Enterococcus faecium L50 ▿

Journal: Applied and Environmental Microbiology

doi: 10.1128/AEM.01476-08

(A) Construction of the S. cerevisiae protein expression and secretion vector pYABD01, derived from pPICZαA and pYES2, containing the yeast gene region encoding the mating pheromone α-factor 1 secretion signal ( MF α 1 s ), including
Figure Legend Snippet: (A) Construction of the S. cerevisiae protein expression and secretion vector pYABD01, derived from pPICZαA and pYES2, containing the yeast gene region encoding the mating pheromone α-factor 1 secretion signal ( MF α 1 s ), including

Techniques Used: Expressing, Plasmid Preparation, Derivative Assay

20) Product Images from "Enhancing the production of cephalosporin C through modulating the autophagic process of Acremonium chrysogenum"

Article Title: Enhancing the production of cephalosporin C through modulating the autophagic process of Acremonium chrysogenum

Journal: Microbial Cell Factories

doi: 10.1186/s12934-018-1021-9

Identification of an ATG8 gene homologue from A. chrysogenum . a Acatg8 with 2 introns. b Sequence alignment of AcAtg8 with its homologs. AcAtg8 shows 96% identity to Atg8 from A. oryzae , 83% identity to Atg8 from U. maydis , 69% identity to Atg8 from D. discoideum , 91% identity to Atg8 from M. oryzae , 78% identity to Atg8 from S. cerevisiae . The asterix indicates the glycine cutting site that is conserved at the C terminal. c Viability of ∆atg8 and its complemented strains. The viability of ∆atg8 and its complemented strains was detected after 18 days of incubation on the nitrogen-starved medium (SG-N). d Distribution of AcAtg8 in A. chrysogenum . Fluorescence observation demonstrated that AcAtg8 was widely distributed throughout the hyphae when the fungal strains grew under nutrient-rich conditions (Nonstarvation), while AcAtg8 was transferred into vacuoles under starvation condition (starvation). WT: the S. cerevisiae wild-type strain; ∆atg8: the S. cerevisiae ATG8 mutant; YC1-3: the complemented strains of ∆atg8; ∆atg8/pYES2: ∆atg8 carrying the plasmid pYES2 as the control; DIC; differential interference contrast; GFP: green fluorescent protein
Figure Legend Snippet: Identification of an ATG8 gene homologue from A. chrysogenum . a Acatg8 with 2 introns. b Sequence alignment of AcAtg8 with its homologs. AcAtg8 shows 96% identity to Atg8 from A. oryzae , 83% identity to Atg8 from U. maydis , 69% identity to Atg8 from D. discoideum , 91% identity to Atg8 from M. oryzae , 78% identity to Atg8 from S. cerevisiae . The asterix indicates the glycine cutting site that is conserved at the C terminal. c Viability of ∆atg8 and its complemented strains. The viability of ∆atg8 and its complemented strains was detected after 18 days of incubation on the nitrogen-starved medium (SG-N). d Distribution of AcAtg8 in A. chrysogenum . Fluorescence observation demonstrated that AcAtg8 was widely distributed throughout the hyphae when the fungal strains grew under nutrient-rich conditions (Nonstarvation), while AcAtg8 was transferred into vacuoles under starvation condition (starvation). WT: the S. cerevisiae wild-type strain; ∆atg8: the S. cerevisiae ATG8 mutant; YC1-3: the complemented strains of ∆atg8; ∆atg8/pYES2: ∆atg8 carrying the plasmid pYES2 as the control; DIC; differential interference contrast; GFP: green fluorescent protein

Techniques Used: Sequencing, Incubation, Fluorescence, Mutagenesis, Plasmid Preparation

21) Product Images from "Identification of a Calmodulin-Regulated Soybean Ca2+-ATPase (SCA1) That Is Located in the Plasma Membrane"

Article Title: Identification of a Calmodulin-Regulated Soybean Ca2+-ATPase (SCA1) That Is Located in the Plasma Membrane

Journal: The Plant Cell

doi:

Complementation of Yeast Mutant K616 through Expression of SCA1p nt85. (A) Complementation of yeast Ca 2+ -ATPases mutant by an N-terminal–truncated mutant pump (SCA1p nt85). Wild-type (W303-1A) and triple mutant (K616) cells were transformed with a vector (pYES2) alone, or with pYES2-SCA1 and pYES2-SCA1 nt85. The cells were streaked onto SC-URA plates containing 10 mM EGTA, pH 6.0, and either glucose (Glc) or galactose (Gal); the cells were incubated for 4 days at 30°C. A diagram indicates yeast strains and transformed vectors. Full-length and ΔN indicate SCA1p and SCA1p nt85, respectively. (B) Sucrose gradient fractionation of SCA1p and SCA1p nt85. Microsomal membranes isolated from yeasts transformed with pYES2-SCA1 and pYES2-SCA1 nt85 were fractionated over a continuous sucrose gradient of 20 to 60% (w/w), and 1-mL fractions were collected from the top of each gradient. Samples (10 μL) from each gradient fraction were subjected to 8% SDS-PAGE and transferred to Immobilon-P membranes. The gel blots were probed with purified anti-SCA1 antibody. Immunoreactive bands were detected by electrochemiluminescence and exposure to x-ray film. Asterisks indicate the peak fractions for plasma membrane fractions, as determined by assays for vanadate-sensitive (calcium-independent) H + -ATPase activity.
Figure Legend Snippet: Complementation of Yeast Mutant K616 through Expression of SCA1p nt85. (A) Complementation of yeast Ca 2+ -ATPases mutant by an N-terminal–truncated mutant pump (SCA1p nt85). Wild-type (W303-1A) and triple mutant (K616) cells were transformed with a vector (pYES2) alone, or with pYES2-SCA1 and pYES2-SCA1 nt85. The cells were streaked onto SC-URA plates containing 10 mM EGTA, pH 6.0, and either glucose (Glc) or galactose (Gal); the cells were incubated for 4 days at 30°C. A diagram indicates yeast strains and transformed vectors. Full-length and ΔN indicate SCA1p and SCA1p nt85, respectively. (B) Sucrose gradient fractionation of SCA1p and SCA1p nt85. Microsomal membranes isolated from yeasts transformed with pYES2-SCA1 and pYES2-SCA1 nt85 were fractionated over a continuous sucrose gradient of 20 to 60% (w/w), and 1-mL fractions were collected from the top of each gradient. Samples (10 μL) from each gradient fraction were subjected to 8% SDS-PAGE and transferred to Immobilon-P membranes. The gel blots were probed with purified anti-SCA1 antibody. Immunoreactive bands were detected by electrochemiluminescence and exposure to x-ray film. Asterisks indicate the peak fractions for plasma membrane fractions, as determined by assays for vanadate-sensitive (calcium-independent) H + -ATPase activity.

Techniques Used: Mutagenesis, Expressing, Transformation Assay, Plasmid Preparation, Gas Chromatography, Incubation, Fractionation, Isolation, SDS Page, Purification, Electrochemiluminescence, Activity Assay

22) Product Images from "Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard"

Article Title: Mutations in the promoter, intron and CDS of two FAD2 generate multiple alleles modulating linoleic acid level in yellow mustard

Journal: Scientific Reports

doi: 10.1038/s41598-017-08317-y

Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.
Figure Legend Snippet: Heterologous expression of the SalFAD2 . LIA 1 alleles LIA 1a , LIA 1b and lia 1 , and SalFAD 2. LIA 2 alleles LIA 2 and lia 2 in Yeast. Gas chromatography analysis of fatty acid composition of yeast cells containing the construct pYES2.1/V5-His-TOPO-LIA 1a with LIA 1 a coding DNA sequence (CDS), pYES2.1/V5-His-TOPO-LIA 1b with LIA 1b CDS, pYES2.1/V5-His-TOPO-lia 1 with lia 1 CDS, pYES2.1/V5-His-TOPO-LIA 2 with LIA 2 CDS, pYES2.1/V5-His-TOPO- lia 2 with lia 2 CDS and the empty vector pYES2.1/V5-His-TOPO. **Statistical significant difference at p = 0.01 level.

Techniques Used: Expressing, Gas Chromatography, Construct, Sequencing, Plasmid Preparation

23) Product Images from "The Cyclase-Associated Protein Cap1 Is Important for Proper Regulation of Infection-Related Morphogenesis in Magnaporthe oryzae"

Article Title: The Cyclase-Associated Protein Cap1 Is Important for Proper Regulation of Infection-Related Morphogenesis in Magnaporthe oryzae

Journal: PLoS Pathogens

doi: 10.1371/journal.ppat.1002911

Assays for the interaction of CAP1 with MAC1 and its function in yeast. A. The domain structure of M. oryzae Cap1, S. cerevisiae Srv2, and human Cap1(hCap1). ACB, AC-binding domain ; P1 and P2, proline-rich regions; AB; actin-binding domain. B. Yeast transformants expressing the CAP1 prey and MAC1 CT bait constructs were assayed for growth on SD-Trp-Leu and SD-His plates and ß-galactosidase activities (LacZ). The Mst11-Mst50 and Pmk1-Mst50 interactions were the positive and negative controls. C. Co-IP assays. Western blots of total proteins and proteins eluted from anti-FLAG M2 beads from transformant CMT ( CAP1 -GFP and MAC1 CT -3×FLAG) and transformant MCF ( MAC1 CT -3×FLAG) were detected with anti-FLAG or anti-GFP antibodies. D. Yeast cells (10 3 to 10 6 cells/ml) of BY4741, Δ srv2 mutant, and Δ srv2 - CAP1 or Δ srv2 -pYES2 transformants were assayed for growth on YPGal (galactose) plates with or without 5 mM H 2 O 2 or 1 M NaCl.
Figure Legend Snippet: Assays for the interaction of CAP1 with MAC1 and its function in yeast. A. The domain structure of M. oryzae Cap1, S. cerevisiae Srv2, and human Cap1(hCap1). ACB, AC-binding domain ; P1 and P2, proline-rich regions; AB; actin-binding domain. B. Yeast transformants expressing the CAP1 prey and MAC1 CT bait constructs were assayed for growth on SD-Trp-Leu and SD-His plates and ß-galactosidase activities (LacZ). The Mst11-Mst50 and Pmk1-Mst50 interactions were the positive and negative controls. C. Co-IP assays. Western blots of total proteins and proteins eluted from anti-FLAG M2 beads from transformant CMT ( CAP1 -GFP and MAC1 CT -3×FLAG) and transformant MCF ( MAC1 CT -3×FLAG) were detected with anti-FLAG or anti-GFP antibodies. D. Yeast cells (10 3 to 10 6 cells/ml) of BY4741, Δ srv2 mutant, and Δ srv2 - CAP1 or Δ srv2 -pYES2 transformants were assayed for growth on YPGal (galactose) plates with or without 5 mM H 2 O 2 or 1 M NaCl.

Techniques Used: Binding Assay, Expressing, Construct, Co-Immunoprecipitation Assay, Western Blot, Mutagenesis

24) Product Images from "Study of model systems to test the potential function of Artemia group 1 late embryogenesis abundant (LEA) proteins"

Article Title: Study of model systems to test the potential function of Artemia group 1 late embryogenesis abundant (LEA) proteins

Journal: Cell Stress & Chaperones

doi: 10.1007/s12192-015-0647-3

Growth of S. cerevisiae doubly transfected with cytoplasmic and mitochondrial Artemia LEA-1 genes. Yeast were transfected with pYES2/Afr-cLEA-1 and pYES3/Afr-mLEA-1 or with pYES2 and pYES3 as controls and grown in SC (w) medium at 30 °C
Figure Legend Snippet: Growth of S. cerevisiae doubly transfected with cytoplasmic and mitochondrial Artemia LEA-1 genes. Yeast were transfected with pYES2/Afr-cLEA-1 and pYES3/Afr-mLEA-1 or with pYES2 and pYES3 as controls and grown in SC (w) medium at 30 °C

Techniques Used: Transfection

Composite of three experiments to determine the effect of three osmoticums on growth of S. cerevisiae transfected with pYES2/pYES3 (control) or pYES2/Afr-cLEA-1 and pYES3/Afr-mLEA-1 (experimental). Control and transfected cells were grown overnight in
Figure Legend Snippet: Composite of three experiments to determine the effect of three osmoticums on growth of S. cerevisiae transfected with pYES2/pYES3 (control) or pYES2/Afr-cLEA-1 and pYES3/Afr-mLEA-1 (experimental). Control and transfected cells were grown overnight in

Techniques Used: Transfection

25) Product Images from "SUMO-independent in vivo activity of a SUMO-targeted ubiquitin ligase toward a short-lived transcription factor"

Article Title: SUMO-independent in vivo activity of a SUMO-targeted ubiquitin ligase toward a short-lived transcription factor

Journal: Genes & Development

doi: 10.1101/gad.1906510

Slx5, but not Slx8, interacts physically with α2. Yeast ubc4Δ ubc6Δ matα2Δ cells (MHY3765) cells were cotransformed with pRS425-GAL1-α2 and a pYES2.1 (2 μm, GAL1 ) plasmid expressing V5-His 6 -tagged
Figure Legend Snippet: Slx5, but not Slx8, interacts physically with α2. Yeast ubc4Δ ubc6Δ matα2Δ cells (MHY3765) cells were cotransformed with pRS425-GAL1-α2 and a pYES2.1 (2 μm, GAL1 ) plasmid expressing V5-His 6 -tagged

Techniques Used: Plasmid Preparation, Expressing

26) Product Images from "A Role For Lte1p (a Low Temperature Essential Protein Involved in Mitosis) in Proprotein Processing in the Yeast Secretory Pathway"

Article Title: A Role For Lte1p (a Low Temperature Essential Protein Involved in Mitosis) in Proprotein Processing in the Yeast Secretory Pathway

Journal: The Journal of biological chemistry

doi: 10.1074/jbc.M610500200

Low temperature growth defect of lte1 1012 and lte1 null ( lte1 Δ) cells is rescued by expression of mycLte1p A , wild-type ( wt ) and lte1 1012 cells were transformed with the empty pYES2 high copy expression plasmid; lte1 1012 mutant cells could not grow efficiently at 13 °C on any carbon source. However, lte1 1012 cells expressing mycLte1p from the GAL1 promoter in pYES2 grew normally on galactose (in which the GAL1 promoter is strongly stimulated) or on raffinose (in which the GAL1 promoter is less strongly stimulated). B , quantitative growth assay on galactose plates in which wild-type ( wt ) yeast growth was compared with lte1 Δ yeast, or lte1 Δ bearing full-length mycLte1p ( third line ) or mycLte1p 1012 ( bottom line ). Each line contains an identical volume of yeast starting at an identical OD, spotted in sequential 10-fold serial dilutions from left to right .
Figure Legend Snippet: Low temperature growth defect of lte1 1012 and lte1 null ( lte1 Δ) cells is rescued by expression of mycLte1p A , wild-type ( wt ) and lte1 1012 cells were transformed with the empty pYES2 high copy expression plasmid; lte1 1012 mutant cells could not grow efficiently at 13 °C on any carbon source. However, lte1 1012 cells expressing mycLte1p from the GAL1 promoter in pYES2 grew normally on galactose (in which the GAL1 promoter is strongly stimulated) or on raffinose (in which the GAL1 promoter is less strongly stimulated). B , quantitative growth assay on galactose plates in which wild-type ( wt ) yeast growth was compared with lte1 Δ yeast, or lte1 Δ bearing full-length mycLte1p ( third line ) or mycLte1p 1012 ( bottom line ). Each line contains an identical volume of yeast starting at an identical OD, spotted in sequential 10-fold serial dilutions from left to right .

Techniques Used: Expressing, Transformation Assay, Plasmid Preparation, Mutagenesis, Growth Assay

27) Product Images from "A docking site determining specificity of Pbs2 MAPKK for Ssk2/Ssk22 MAPKKKs in the yeast HOG pathway"

Article Title: A docking site determining specificity of Pbs2 MAPKK for Ssk2/Ssk22 MAPKKKs in the yeast HOG pathway

Journal: The EMBO Journal

doi: 10.1093/emboj/cdg353

Fig. 5. ‘Cross-talk’ activation of the mating pathway by Ssk2/Ssk22 via binding to a Pbs2–Ste7 fusion protein. ( A ) Schematic diagram of the Pbs2–Ste7 fusion protein used in this analysis. ( B ) Induction of FUS1-lacZ expression following activation of the Pbs2–Ste7 fusion protein by constitutively active Ssk2ΔN or Ssk22ΔN. The reporter strain KT007 ( pbs2 Δ FUS1::lacZ::LEU2 ) was co-transformed with either pYES2 (Vector), pYES2-Ssk2ΔN or pYES2-Ssk22ΔN together with either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in SRaf medium, and were either harvested (–Gal) or incubated further for 2 h in the presence of 2.5% galactose (+Gal). FUS1-lacZ expression was measured by assaying the β-galactosidase activities in cell lysates as described in Materials and methods. For each combination of plasmids, three independent transformants were assayed in triplicate, and the average activity ± SD is shown. ( C ) FUS1-lacZ expression induced by osmotic stress via the Pbs2–Ste7 fusion protein. KT005 ( pbs2 Δ ste11 Δ) was co-transformed with a FUS1-lacZ reporter plasmid, pSB231, and either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in CAD and either harvested (–NaCl), or incubated further for 4 h following the addition of 0.4 M NaCl (+NaCl). ( D ) Restoration of pheromone-induced FUS1-lacZ expression in the ste7 Δ mutant by expression of the Pbs2–Ste7 or Pbs2 V54G –Ste7 hybrid protein. FP56 ( ste7 Δ) was co-transformed with pSB231, and either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in CAD and either harvested (–αF), or incubated further for 2 h following the addition of 5 µM α-factor (+αF). FUS1-lacZ expression was measured as in (B).
Figure Legend Snippet: Fig. 5. ‘Cross-talk’ activation of the mating pathway by Ssk2/Ssk22 via binding to a Pbs2–Ste7 fusion protein. ( A ) Schematic diagram of the Pbs2–Ste7 fusion protein used in this analysis. ( B ) Induction of FUS1-lacZ expression following activation of the Pbs2–Ste7 fusion protein by constitutively active Ssk2ΔN or Ssk22ΔN. The reporter strain KT007 ( pbs2 Δ FUS1::lacZ::LEU2 ) was co-transformed with either pYES2 (Vector), pYES2-Ssk2ΔN or pYES2-Ssk22ΔN together with either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in SRaf medium, and were either harvested (–Gal) or incubated further for 2 h in the presence of 2.5% galactose (+Gal). FUS1-lacZ expression was measured by assaying the β-galactosidase activities in cell lysates as described in Materials and methods. For each combination of plasmids, three independent transformants were assayed in triplicate, and the average activity ± SD is shown. ( C ) FUS1-lacZ expression induced by osmotic stress via the Pbs2–Ste7 fusion protein. KT005 ( pbs2 Δ ste11 Δ) was co-transformed with a FUS1-lacZ reporter plasmid, pSB231, and either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in CAD and either harvested (–NaCl), or incubated further for 4 h following the addition of 0.4 M NaCl (+NaCl). ( D ) Restoration of pheromone-induced FUS1-lacZ expression in the ste7 Δ mutant by expression of the Pbs2–Ste7 or Pbs2 V54G –Ste7 hybrid protein. FP56 ( ste7 Δ) was co-transformed with pSB231, and either YCplac22′ (Vector), YCplac22′-Pbs2–Ste7 or YCplac22′-Pbs2 V54G –Ste7. The cells were grown in CAD and either harvested (–αF), or incubated further for 2 h following the addition of 5 µM α-factor (+αF). FUS1-lacZ expression was measured as in (B).

Techniques Used: Activation Assay, Binding Assay, Expressing, Transformation Assay, Plasmid Preparation, Incubation, Activity Assay, Mutagenesis

28) Product Images from "Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions"

Article Title: Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions

Journal: Nucleic Acids Research

doi: 10.1093/nar/gkv695

Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.
Figure Legend Snippet: Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.

Techniques Used: Functional Assay, Sequencing, Expressing

29) Product Images from "Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption"

Article Title: Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption

Journal: International Journal for Parasitology: Drugs and Drug Resistance

doi: 10.1016/j.ijpddr.2018.06.003

Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.
Figure Legend Snippet: Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.

Techniques Used: Functional Assay, Mutagenesis, Transformation Assay, Expressing, Plasmid Preparation, Positive Control

30) Product Images from "HSP101 functions as a specific translational regulatory protein whose activity is regulated by nutrient status"

Article Title: HSP101 functions as a specific translational regulatory protein whose activity is regulated by nutrient status

Journal: Genes & Development

doi:

p102 from tobacco and wheat is a functional HSP101 that complements a thermotolerance defect in yeast. ( A ) SL304A, the hsp104 mutant containing p GAL1–NtHSP101 or p GAL1–TaHSP101 , was grown to an early exponential stage in SGM (an OD of 0.06 or 1.8 × 10 6 cells/ml) prior to assaying for thermotolerance. The expression vector, pYES2, was used as a negative control and pYS104, containing HSP104 , was used as a positive control. The percentage of survival at 50°C was plotted as a function of the length of treatment. ( B ) SL304A(p GAL1–NtHSP101 ) was grown in galactose, glucose, or raffinose prior to assaying for thermotolerance. SL304A(pYES2) grown in galactose was included as a negative control. ( C ) Thermotolerance conferred by tobacco HSP101 in late-exponential yeast (an OD of 1.2 or 3.6 × 10 7 cells/ml) was examined.
Figure Legend Snippet: p102 from tobacco and wheat is a functional HSP101 that complements a thermotolerance defect in yeast. ( A ) SL304A, the hsp104 mutant containing p GAL1–NtHSP101 or p GAL1–TaHSP101 , was grown to an early exponential stage in SGM (an OD of 0.06 or 1.8 × 10 6 cells/ml) prior to assaying for thermotolerance. The expression vector, pYES2, was used as a negative control and pYS104, containing HSP104 , was used as a positive control. The percentage of survival at 50°C was plotted as a function of the length of treatment. ( B ) SL304A(p GAL1–NtHSP101 ) was grown in galactose, glucose, or raffinose prior to assaying for thermotolerance. SL304A(pYES2) grown in galactose was included as a negative control. ( C ) Thermotolerance conferred by tobacco HSP101 in late-exponential yeast (an OD of 1.2 or 3.6 × 10 7 cells/ml) was examined.

Techniques Used: Functional Assay, Mutagenesis, Expressing, Plasmid Preparation, Negative Control, Positive Control

NtHSP101 specifically enhancesexpression from an mRNA when Ω is present as the 5′ leader. ( A ) Expression from p TPI –Ω- luc (•) or p TPI–luc (○) was followed in SL304A transformed with p GAL1–NtHSP101 ( top and middle ) or pYES2 ( bottom ) to examine the regulatory role of NtHSP101 during translation. Transformants were first grown to late-exponential stage (whereupon expression from p TPI –Ω- luc and p TPI–luc was equivalent) and then inoculated into synthetic galactose medium ( top and bottom ) or synthetic dextrose medium ( middle ) at the zero time point. Luciferase expression was measured at time points during the growth cycle, normalized to the OD ( right ) during growth. The expression from p GAL1 –Ω- luc , p GAL1–luc , and their expression ratio (i.e., NtHSP101 activity as indicated by the ratio, Ω- luc/luc ) are shown below each panel. ( B ) Tobacco and wheat HSP101 retain RNA-binding activity when expressed in yeast. ( Top ) SL304A containing p GAL1–NtHSP101 or p GAL1–TaHSP101 were grown in galactose or glucose and crude extracts used in RNA gel shift binding assays with radiolabeled Ω RNA. SL304A(pYES2) was used as a negative control and purified HSP101 was included as a positive control. ( Middle ) Western analysis of crude extracts from SL304A (containing either p GAL1–NtHSP101 , p GAL1–TaHSP101 , or pYES2) by use of anti-p102 (i.e., anti-HSP101) antibodies. The lanes correspond to the extracts as indicated at top . The lower molecular weight bands in lane 1 are degradation products of HSP101 that are observed when a high level of the purified protein is analyzed. ( Bottom ) Titration of HSP101 protein in the Ω RNA gel shift binding assay. The concentration of purified HSP101 used in each binding reaction was decreased twofold in each succeeding lane, whereas the concentration of Ω RNA was held constant.
Figure Legend Snippet: NtHSP101 specifically enhancesexpression from an mRNA when Ω is present as the 5′ leader. ( A ) Expression from p TPI –Ω- luc (•) or p TPI–luc (○) was followed in SL304A transformed with p GAL1–NtHSP101 ( top and middle ) or pYES2 ( bottom ) to examine the regulatory role of NtHSP101 during translation. Transformants were first grown to late-exponential stage (whereupon expression from p TPI –Ω- luc and p TPI–luc was equivalent) and then inoculated into synthetic galactose medium ( top and bottom ) or synthetic dextrose medium ( middle ) at the zero time point. Luciferase expression was measured at time points during the growth cycle, normalized to the OD ( right ) during growth. The expression from p GAL1 –Ω- luc , p GAL1–luc , and their expression ratio (i.e., NtHSP101 activity as indicated by the ratio, Ω- luc/luc ) are shown below each panel. ( B ) Tobacco and wheat HSP101 retain RNA-binding activity when expressed in yeast. ( Top ) SL304A containing p GAL1–NtHSP101 or p GAL1–TaHSP101 were grown in galactose or glucose and crude extracts used in RNA gel shift binding assays with radiolabeled Ω RNA. SL304A(pYES2) was used as a negative control and purified HSP101 was included as a positive control. ( Middle ) Western analysis of crude extracts from SL304A (containing either p GAL1–NtHSP101 , p GAL1–TaHSP101 , or pYES2) by use of anti-p102 (i.e., anti-HSP101) antibodies. The lanes correspond to the extracts as indicated at top . The lower molecular weight bands in lane 1 are degradation products of HSP101 that are observed when a high level of the purified protein is analyzed. ( Bottom ) Titration of HSP101 protein in the Ω RNA gel shift binding assay. The concentration of purified HSP101 used in each binding reaction was decreased twofold in each succeeding lane, whereas the concentration of Ω RNA was held constant.

Techniques Used: Expressing, Transformation Assay, Luciferase, Activity Assay, RNA Binding Assay, Electrophoretic Mobility Shift Assay, Binding Assay, Negative Control, Purification, Positive Control, Western Blot, Molecular Weight, Titration, Concentration Assay

HSP101 enhancement is specific to Ω-containing mRNAs and is not dependent on a poly(A) tail. ( A ) In vitro-synthesized luc mRNA constructs terminating with or without a poly(A) tail or with the TMV 3′UTR (indicated in each panel) were electroporated into SL304A (containing either p GAL1–NtHSP101 or pYES2 as indicated at left ) and expression measured following the completion of translation. The expression ratio is shown at right . ( B ) Ω- luc -A 50 and luc -A 50 mRNAs were introduced into SGM-grown, SL304A(p GAL1–NtHSP101 ), SL304A(p GAL1–TaHSP101 ), or SL304A(pYES2) by electroporation and expression measured following the completion of translation. The expression ratio is shown at right . ( C ) NtHSP101 increases the rate of translation from an Ω-containing mRNA. Ω- luc -A 50 and luc -A 50 mRNAs were introduced into SGM-grown, SL304A(p GAL1–NtHSP101 ) cells by electroporation, luciferase expression measured following RNA delivery, and the data plotted as a function of the time of translation. ( D ) NtHSP101 does not regulate translation from TEV 5′ leader-containing mRNA. TEV- luc -A 50 , Ω- luc -A 50 , or luc -A 50 mRNAs were electroporated into SL304A containing either p GAL1–NtHSP101 or pYES2. Expression is shown relative to the level obtained from each construct in SL304A(pYES2).
Figure Legend Snippet: HSP101 enhancement is specific to Ω-containing mRNAs and is not dependent on a poly(A) tail. ( A ) In vitro-synthesized luc mRNA constructs terminating with or without a poly(A) tail or with the TMV 3′UTR (indicated in each panel) were electroporated into SL304A (containing either p GAL1–NtHSP101 or pYES2 as indicated at left ) and expression measured following the completion of translation. The expression ratio is shown at right . ( B ) Ω- luc -A 50 and luc -A 50 mRNAs were introduced into SGM-grown, SL304A(p GAL1–NtHSP101 ), SL304A(p GAL1–TaHSP101 ), or SL304A(pYES2) by electroporation and expression measured following the completion of translation. The expression ratio is shown at right . ( C ) NtHSP101 increases the rate of translation from an Ω-containing mRNA. Ω- luc -A 50 and luc -A 50 mRNAs were introduced into SGM-grown, SL304A(p GAL1–NtHSP101 ) cells by electroporation, luciferase expression measured following RNA delivery, and the data plotted as a function of the time of translation. ( D ) NtHSP101 does not regulate translation from TEV 5′ leader-containing mRNA. TEV- luc -A 50 , Ω- luc -A 50 , or luc -A 50 mRNAs were electroporated into SL304A containing either p GAL1–NtHSP101 or pYES2. Expression is shown relative to the level obtained from each construct in SL304A(pYES2).

Techniques Used: In Vitro, Synthesized, Construct, Expressing, Electroporation, Luciferase

The NtHSP101-mediated regulation is not affected following the inactivation of eIF4A in the conditional mutant, SS8-3B(pSSC120), or in yeast expressing eIF2α mutants affecting factor activity. Translation from in vitro-synthesized capped Ω- luc -A 50 and luc -A 50 mRNAs was measured following RNA delivery into early exponential ( A ) wild-type CW04 and eIF4A ts mutant cells grown at the permissive (30°C) or nonpermissive (37°C) temperatures or ( B ) gcn2 containing wild-type eIF2α, i.e., gcn2 (eIF2α-WT), GCN2 containing wild-type eIF2α, i.e., GCN2 (eIF2α-WT), GCN2 containing the eIF2α-S51A mutant, or GCN2 containing the eIF2α-S51D mutant, each containing either p GAL1–NtHSP101 or pYES2. The expression ratio is shown to the right of each pair of histograms. (Solid bars) Ω- luc -A 50 ; (hatched bars) luc -A 50 .
Figure Legend Snippet: The NtHSP101-mediated regulation is not affected following the inactivation of eIF4A in the conditional mutant, SS8-3B(pSSC120), or in yeast expressing eIF2α mutants affecting factor activity. Translation from in vitro-synthesized capped Ω- luc -A 50 and luc -A 50 mRNAs was measured following RNA delivery into early exponential ( A ) wild-type CW04 and eIF4A ts mutant cells grown at the permissive (30°C) or nonpermissive (37°C) temperatures or ( B ) gcn2 containing wild-type eIF2α, i.e., gcn2 (eIF2α-WT), GCN2 containing wild-type eIF2α, i.e., GCN2 (eIF2α-WT), GCN2 containing the eIF2α-S51A mutant, or GCN2 containing the eIF2α-S51D mutant, each containing either p GAL1–NtHSP101 or pYES2. The expression ratio is shown to the right of each pair of histograms. (Solid bars) Ω- luc -A 50 ; (hatched bars) luc -A 50 .

Techniques Used: Mutagenesis, Expressing, Activity Assay, In Vitro, Synthesized

31) Product Images from "The Thellungiella salsuginea Tonoplast Aquaporin TsTIP1;2 Functions in Protection Against Multiple Abiotic Stresses"

Article Title: The Thellungiella salsuginea Tonoplast Aquaporin TsTIP1;2 Functions in Protection Against Multiple Abiotic Stresses

Journal: Plant and Cell Physiology

doi: 10.1093/pcp/pct166

H 2 O 2 permeability of yeast cells expressing TsTIP1;2 . (A) Saccharomyces cerevisiae aqy-null strain cells transformed with the empty vector pYES2 alone or pYES2-TsTIP1;2 were spotted in 10-fold dilutions on medium without or with 1, 1.5 or 2 mM H 2 O 2 , respectively.
Figure Legend Snippet: H 2 O 2 permeability of yeast cells expressing TsTIP1;2 . (A) Saccharomyces cerevisiae aqy-null strain cells transformed with the empty vector pYES2 alone or pYES2-TsTIP1;2 were spotted in 10-fold dilutions on medium without or with 1, 1.5 or 2 mM H 2 O 2 , respectively.

Techniques Used: Permeability, Expressing, Transformation Assay, Plasmid Preparation

32) Product Images from "Functional Characterization and Localization of Pneumocystis carinii Lanosterol Synthase ▿ Lanosterol Synthase ▿ †"

Article Title: Functional Characterization and Localization of Pneumocystis carinii Lanosterol Synthase ▿ Lanosterol Synthase ▿ †

Journal: Eukaryotic Cell

doi: 10.1128/EC.00264-09

PcErg7p localizes to lipid particles in yeast. Lipid particles were isolated from wild-type yeast and yeast containing pYES2.1, pYES2.1/Pc ERG7 , or pYES2.1/Sc ERG7 . The floating layer was removed in 1-ml aliquots, and 5-μg portions of protein from
Figure Legend Snippet: PcErg7p localizes to lipid particles in yeast. Lipid particles were isolated from wild-type yeast and yeast containing pYES2.1, pYES2.1/Pc ERG7 , or pYES2.1/Sc ERG7 . The floating layer was removed in 1-ml aliquots, and 5-μg portions of protein from

Techniques Used: Isolation

Detection of wild-type ScErg7p, recombinant PcErg7p and ScErg7p. Protein extracts from P. carinii , S. cerevisiae , and S. cerevisiae containing either pYES2.1/Pc ERG7 or pYES2.1/Sc ERG7 , were blotted and probed with PcErg7p antiserum. Lanes 1 to 5 correspond
Figure Legend Snippet: Detection of wild-type ScErg7p, recombinant PcErg7p and ScErg7p. Protein extracts from P. carinii , S. cerevisiae , and S. cerevisiae containing either pYES2.1/Pc ERG7 or pYES2.1/Sc ERG7 , were blotted and probed with PcErg7p antiserum. Lanes 1 to 5 correspond

Techniques Used: Recombinant

Growth curve comparing growth of wild-type yeast (WT) and yeast containing, pYES2.1 (EV), pYES2.1/Pc ERG7 (Pc), or pYES2.1/Sc ERG7 (Sc) cultured in liquid medium at 30°C. Each data point represents the mean of three independent studies. Error bars
Figure Legend Snippet: Growth curve comparing growth of wild-type yeast (WT) and yeast containing, pYES2.1 (EV), pYES2.1/Pc ERG7 (Pc), or pYES2.1/Sc ERG7 (Sc) cultured in liquid medium at 30°C. Each data point represents the mean of three independent studies. Error bars

Techniques Used: Cell Culture

Lanosterol production by wild-type yeast (WT) or yeast containing either pYES2.1/Sc ERG7 (SC) or pYES2.1/Pc ERG7 (PC). Lanosterol levels were assessed by gas liquid chromatography, and asterisks indicate statistical significance. Values represent the mean
Figure Legend Snippet: Lanosterol production by wild-type yeast (WT) or yeast containing either pYES2.1/Sc ERG7 (SC) or pYES2.1/Pc ERG7 (PC). Lanosterol levels were assessed by gas liquid chromatography, and asterisks indicate statistical significance. Values represent the mean

Techniques Used: Gas Chromatography

33) Product Images from "Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions"

Article Title: Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions

Journal: Nucleic Acids Research

doi: 10.1093/nar/gkv695

Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.
Figure Legend Snippet: Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.

Techniques Used: Functional Assay, Sequencing, Expressing

34) Product Images from "A Conserved Non-Canonical Docking Mechanism Regulates the Binding of Dual Specificity Phosphatases to Cell Integrity Mitogen-Activated Protein Kinases (MAPKs) in Budding and Fission Yeasts"

Article Title: A Conserved Non-Canonical Docking Mechanism Regulates the Binding of Dual Specificity Phosphatases to Cell Integrity Mitogen-Activated Protein Kinases (MAPKs) in Budding and Fission Yeasts

Journal: PLoS ONE

doi: 10.1371/journal.pone.0085390

Effect of the lack of the IYT motif of Sdp1 on the CWI pathway. (A) Western blotting analysis of the Y00897 ( gas1 Δ) strain transformed with pYES2 (vector), pYES2SDP1myc (Sdp1), pYES2SDP1 IAYATA myc (Sdp1 IAYATA ), pYES2SDP1 C140A (Sdp1 C140A ) or pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ) plasmids. Cells were cultured to mid-log phase in raffinose-based selective medium at 24°C, then galactose was added to a final 2% concentration for 4 hours at 24°C, and protein extracts were prepared. Phospho-Slt2, Sdp1-Myc and G6PDH (as a loading control) were detected with anti-phospho-p42/44 (upper panel), anti-Myc (middle panel) and anti-G6PDH (lower panel) antibodies, respectively. (B) Expression of MLP1-lacZ was examined by determining β-galactosidase activity in cell extracts of the same transformants as in A also carrying the pMLP1 -LACZ plasmid. Data shown are the average of three independent experiments performed in duplicate. Error bars indicate standard deviations. (C) Western blotting analysis of the wild type YPH499 strain transformed with the same plasmids as in Figure 2A . Cell growth conditions, extract preparation and immunoblot analysis was performed as in Figure 2A . (D) In vivo localisation of Slt2-GFP in YPH499 yeast cells carrying the plasmid pRS425- SLT2 -GFP transformed with pYES2 (vector), pYES2SDP1 C140A (Sdp1 C140A ) or pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ) plasmids. Cells were grown to mid-log phase in raffinose-based selective medium at 24°C, and then galactose was added to a final 2% concentration for 4 hours at 24°C. Fluorescence microscopy photographs showing the cellular distribution of the fluorescent Slt2-GFP were taken. Bars, 5 µm. The percentage of cells showing nuclear accumulation of Slt2-GFP from the same cultures was determined. Data shown are the average of three independent experiments in which n > 100. Error bars indicate standard deviations (E) Western blotting analysis of the WT BY4741 strain transformed with pYES2 (vector), pYES2SDP1 C140A (Sdp1 C140A ), pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ), pYES2SDP1 C140A-IA (Sdp1 C140A-IA ), pYES2SDP1 C140A-YA (Sdp1 C140A-YA ) or pYES2SDP1 C140A-TA (Sdp1 C140A-TA ) plasmids. Cells were cultured and extracts processed as in Figure 3A . Similar results were obtained in three different experiments and selected images correspond to representative blots.
Figure Legend Snippet: Effect of the lack of the IYT motif of Sdp1 on the CWI pathway. (A) Western blotting analysis of the Y00897 ( gas1 Δ) strain transformed with pYES2 (vector), pYES2SDP1myc (Sdp1), pYES2SDP1 IAYATA myc (Sdp1 IAYATA ), pYES2SDP1 C140A (Sdp1 C140A ) or pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ) plasmids. Cells were cultured to mid-log phase in raffinose-based selective medium at 24°C, then galactose was added to a final 2% concentration for 4 hours at 24°C, and protein extracts were prepared. Phospho-Slt2, Sdp1-Myc and G6PDH (as a loading control) were detected with anti-phospho-p42/44 (upper panel), anti-Myc (middle panel) and anti-G6PDH (lower panel) antibodies, respectively. (B) Expression of MLP1-lacZ was examined by determining β-galactosidase activity in cell extracts of the same transformants as in A also carrying the pMLP1 -LACZ plasmid. Data shown are the average of three independent experiments performed in duplicate. Error bars indicate standard deviations. (C) Western blotting analysis of the wild type YPH499 strain transformed with the same plasmids as in Figure 2A . Cell growth conditions, extract preparation and immunoblot analysis was performed as in Figure 2A . (D) In vivo localisation of Slt2-GFP in YPH499 yeast cells carrying the plasmid pRS425- SLT2 -GFP transformed with pYES2 (vector), pYES2SDP1 C140A (Sdp1 C140A ) or pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ) plasmids. Cells were grown to mid-log phase in raffinose-based selective medium at 24°C, and then galactose was added to a final 2% concentration for 4 hours at 24°C. Fluorescence microscopy photographs showing the cellular distribution of the fluorescent Slt2-GFP were taken. Bars, 5 µm. The percentage of cells showing nuclear accumulation of Slt2-GFP from the same cultures was determined. Data shown are the average of three independent experiments in which n > 100. Error bars indicate standard deviations (E) Western blotting analysis of the WT BY4741 strain transformed with pYES2 (vector), pYES2SDP1 C140A (Sdp1 C140A ), pYES2SDP1 C140A-IAYATA (Sdp1 C140A-IAYATA ), pYES2SDP1 C140A-IA (Sdp1 C140A-IA ), pYES2SDP1 C140A-YA (Sdp1 C140A-YA ) or pYES2SDP1 C140A-TA (Sdp1 C140A-TA ) plasmids. Cells were cultured and extracts processed as in Figure 3A . Similar results were obtained in three different experiments and selected images correspond to representative blots.

Techniques Used: Western Blot, Transformation Assay, Plasmid Preparation, Cell Culture, Concentration Assay, Expressing, Activity Assay, In Vivo, Fluorescence, Microscopy, IA

35) Product Images from "Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption"

Article Title: Characterization of Divalent Metal Transporter 1 (DMT1) in Brugia malayi suggests an intestinal-associated pathway for iron absorption

Journal: International Journal for Parasitology: Drugs and Drug Resistance

doi: 10.1016/j.ijpddr.2018.06.003

Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.
Figure Legend Snippet: Functional analysis of BmDMT1 in yeast. The fet3fet4 mutant yeast strain was transformed with BmDMT1 or the empty PYES2 expression vector. The DY150 parent strain was used as a positive control and transformed with either BmDMT1 or the empty vector. Serially diluted cells were spotted on URA-selective plates supplemented with either A. 100 μM or 200 μM ferrous ammonium sulfate, or B. 2 μM or 20 μM ferric chloride.

Techniques Used: Functional Assay, Mutagenesis, Transformation Assay, Expressing, Plasmid Preparation, Positive Control

36) Product Images from "Improved Cd, Zn and Mn tolerance and reduced Cd accumulation in grains with wheat-based cell number regulator TaCNR2"

Article Title: Improved Cd, Zn and Mn tolerance and reduced Cd accumulation in grains with wheat-based cell number regulator TaCNR2

Journal: Scientific Reports

doi: 10.1038/s41598-018-37352-6

TaCNR2 -transgenic yeast grown under heavy metal treatments. The pYES2 and pYES2- TaCNR2 were transformed into yeast strain (BY4741). Gradient dilution was spotted onto the solid media (yeast extract; peptone; galactose) with 50 μM CdSO 4 , 5 mM ZnSO 4 and 5 mM MnSO 4 . The YPD medium (yeast extract; peptone; glucose) was the control. Growth was maintained at 30 °C for 3–7 days.
Figure Legend Snippet: TaCNR2 -transgenic yeast grown under heavy metal treatments. The pYES2 and pYES2- TaCNR2 were transformed into yeast strain (BY4741). Gradient dilution was spotted onto the solid media (yeast extract; peptone; galactose) with 50 μM CdSO 4 , 5 mM ZnSO 4 and 5 mM MnSO 4 . The YPD medium (yeast extract; peptone; glucose) was the control. Growth was maintained at 30 °C for 3–7 days.

Techniques Used: Transgenic Assay, Transformation Assay

37) Product Images from "Identification of Amino Acids Conferring Chain Length Substrate Specificities on Fatty Alcohol-forming Reductases FAR5 and FAR8 from Arabidopsis thaliana *"

Article Title: Identification of Amino Acids Conferring Chain Length Substrate Specificities on Fatty Alcohol-forming Reductases FAR5 and FAR8 from Arabidopsis thaliana *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M113.499715

Gas chromatograms of internal lipids of yeast expressing Arabidopsis FAR5, FAR8, or FAR8-S363P. The empty vector pYES2-His 6 /T7 tag acted as a negative control. Transformants were cultured in galactose media to induce protein expression. Fatty acids were
Figure Legend Snippet: Gas chromatograms of internal lipids of yeast expressing Arabidopsis FAR5, FAR8, or FAR8-S363P. The empty vector pYES2-His 6 /T7 tag acted as a negative control. Transformants were cultured in galactose media to induce protein expression. Fatty acids were

Techniques Used: Expressing, Plasmid Preparation, Negative Control, Cell Culture

38) Product Images from "A Mitochondrial Autonomously Replicating Sequence from Pichia pastoris for Uniform High Level Recombinant Protein Production"

Article Title: A Mitochondrial Autonomously Replicating Sequence from Pichia pastoris for Uniform High Level Recombinant Protein Production

Journal: Frontiers in Microbiology

doi: 10.3389/fmicb.2017.00780

(A) S. cerevisiae transformation efficiencies of pYES2 (5.9 kb) and pYES2-Mito (6.4 kb). Error bars represent the standard deviation, with n = 3. (B) PCR assay for circular plasmids in pYES2 and pYES2-Mito clones. Lanes: (M) Marker (1–5) PCR product of five pYES2 strains, using the primer pair pYES2_Circ-FW/RV (6–10) PCR product of five pYES2-Mito strains, applying the primer pair pMito_Circ-mtDNA-FW/RV. The size of relevant bands has been highlighted.
Figure Legend Snippet: (A) S. cerevisiae transformation efficiencies of pYES2 (5.9 kb) and pYES2-Mito (6.4 kb). Error bars represent the standard deviation, with n = 3. (B) PCR assay for circular plasmids in pYES2 and pYES2-Mito clones. Lanes: (M) Marker (1–5) PCR product of five pYES2 strains, using the primer pair pYES2_Circ-FW/RV (6–10) PCR product of five pYES2-Mito strains, applying the primer pair pMito_Circ-mtDNA-FW/RV. The size of relevant bands has been highlighted.

Techniques Used: Transformation Assay, Standard Deviation, Polymerase Chain Reaction, Clone Assay, Marker

39) Product Images from "Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions"

Article Title: Ty1 retrovirus-like element Gag contains overlapping restriction factor and nucleic acid chaperone functions

Journal: Nucleic Acids Research

doi: 10.1093/nar/gkv695

Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.
Figure Legend Snippet: Functional organization of GAG and coexpression of subgenomic segments with pGTy1 his3-AI . ( A ) At the top is the mature Gag (p45) coding sequence with selected ATG codons highlighted (green) and below are segments expressed ectopically from the pYES2 GAL1 promoter. At the bottom, XtalPred ( http://ffas.burnham.org/XtalPred-cgi/xtal.pl ) was used to predict Gag disordered (yellow) and α-helical (red) regions. Also shown is an invariant tryptophan residue (W184) found in Pseudoviridae Gag proteins ( 19 ), the position of the nucleic acid chaperone region ( 21 ), a C-terminal disordered region (C-DR), and the Ty1 protease (PR) cleavage site (H401-N402). An ATG codon was added adjacent to P173 for expression of the CTR and sCTR. ( B ) Two plasmids, pGTy1 his3-AI ( TRP1 , CEN) and GAG segment under pYES2 ( URA3 , 2 μ) were induced from the GAL1 promoter in a Ty1-less Saccharomyces paradoxus strain to determine whether different Gag proteins inhibited Ty1 his3-AI mobility (Table 2 ). ( C ) Total cell protein from induced cultures was immunoblotted with p18 antiserum (anti-p18) or TY-tag (anti-TY-tag). Histidyl tRNA synthetase (anti-Hts1) served as a loading control.

Techniques Used: Functional Assay, Sequencing, Expressing

40) Product Images from "Functional Characterization and Expression Analysis of a Gene, OsENT2, Encoding an Equilibrative Nucleoside Transporter in Rice Suggest a Function in Cytokinin Transport 1"

Article Title: Functional Characterization and Expression Analysis of a Gene, OsENT2, Encoding an Equilibrative Nucleoside Transporter in Rice Suggest a Function in Cytokinin Transport 1

Journal:

doi: 10.1104/pp.105.060137

Growth analysis of yeast cells expressing OsENTs. Growth of yeast cells harboring expression plasmids containing OsENT1 , OsENT2 , OsENT3 , OsENT4 , or controls (BY4735; BY4735 plain recipient cells, ΔFUI1; ΔFUI1 plain recipient cells, pYES2;
Figure Legend Snippet: Growth analysis of yeast cells expressing OsENTs. Growth of yeast cells harboring expression plasmids containing OsENT1 , OsENT2 , OsENT3 , OsENT4 , or controls (BY4735; BY4735 plain recipient cells, ΔFUI1; ΔFUI1 plain recipient cells, pYES2;

Techniques Used: Expressing

Related Articles

Clone Assay:

Article Title: A Type 2C Protein Phosphatase FgPtc3 Is Involved in Cell Wall Integrity, Lipid Metabolism, and Virulence in Fusarium graminearum
Article Snippet: .. The PCR product was digested with Hind III and Sac I, cloned into the pYES2 vector (Invitrogen), and transformed into the yeast mutant BY4741ΔPtc1. .. Yeast transformants were selected on synthetic medium lacking Ura (Clontech).

Article Title: Reconstitution of the mammalian PI3K/PTEN/Akt pathway in yeast
Article Snippet: .. Cassettes in which cDNAs encoding either c-Akt, c-AktK179M or an N-myristoylated c-Akt were fused in frame to an HA (haemagglutinin) epitope-tagged version of eGFP (enhanced green fluorescence protein) [HA–eGFP-Akt, HA–eGFP-AktK179M and myr-HA–eGFP-Akt respectively] were excised with HindIII and BamHI from the original Pcefl(X)-derived plasmids that were constructed for expression in mammalian cells [ ] (a gift from M. Lorenzo, Universidad Complutense, Madrid, Spain) and cloned into the corresponding sites in yeast vector pYES2 (Invitrogen), yielding plasmids pYES-GFP-c-Akt, pYES-GFP-c-AktK179M and pYES-myr-GFP-c-Akt respectively. .. The cDNA encoding PTEN was excised with EcoRI from plasmid Pcmvpten [ ] [a gift from J.M.

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Article Title: Cloning and Functional Characterization of a Vacuolar Na+/H+ Antiporter Gene from Mungbean (VrNHX1) and Its Ectopic Expression Enhanced Salt Tolerance in Arabidopsis thaliana
Article Snippet: .. The CDS of VrNHX1 was cloned into yeast expression vector pYES2.0 (Invitrogen, Carlsbad, CA, USA) with restriction sites of KpnI and BamHI. .. The yeast strains were transformed with pYES2.0 empty vector (labeled as AXTYES2.0 strain) or pYESVrNHX1 recombinant plasmid (labeled as AXTVrNHX1 strain) by Lithium acetate method and selected on SC ura− medium.

Article Title: Functional Desaturase Fads1 (Δ5) and Fads2 (Δ6) Orthologues Evolved before the Origin of Jawed Vertebrates
Article Snippet: .. After restriction digest, the insert was cloned into pYES2 yeast expression vector (Invitrogen), and sequenced. .. The constructs pYES2-ScaFads1 and pYES2-ScaFads2 were transformed into Saccharomyces cerevisiae competent cells (strain InvSc, Invitrogen).

Article Title: The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea
Article Snippet: .. The full-length cDNA of BcATG8 was cloned into the yeast expression vector pYES2 (Invitrogen, Carlsbad, CA) and transformed into the yeast ATG8 null mutant using the lithium acetate (LiAc)/single-stranded DNA (ssDNA)/polyethylene glycol (PEG) transformation protocol ( ). .. Yeast transformants were selected on SD medium lacking uracil (Clontech, Palo Alta, CA).

Article Title: Friedelin Synthase from Maytenus ilicifolia: Leucine 482 Plays an Essential Role in the Production of the Most Rearranged Pentacyclic Triterpene
Article Snippet: .. The full length Maytenus ilicifolia OSC ORFs were cloned into the yeast expression vector pYES2 (Invitrogen), which is under the transcriptional control of galactose (GAL1 promoter). .. The pYES2-Mi FRS and pYES2-Mi CAS1- plasmids were transformed into VZL 1303, and the resulting strains were grown in 0.5 L of synthetic complete medium without uracil (SC-U).

Functional Assay:

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Amplification:

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Fluorescence:

Article Title: Reconstitution of the mammalian PI3K/PTEN/Akt pathway in yeast
Article Snippet: .. Cassettes in which cDNAs encoding either c-Akt, c-AktK179M or an N-myristoylated c-Akt were fused in frame to an HA (haemagglutinin) epitope-tagged version of eGFP (enhanced green fluorescence protein) [HA–eGFP-Akt, HA–eGFP-AktK179M and myr-HA–eGFP-Akt respectively] were excised with HindIII and BamHI from the original Pcefl(X)-derived plasmids that were constructed for expression in mammalian cells [ ] (a gift from M. Lorenzo, Universidad Complutense, Madrid, Spain) and cloned into the corresponding sites in yeast vector pYES2 (Invitrogen), yielding plasmids pYES-GFP-c-Akt, pYES-GFP-c-AktK179M and pYES-myr-GFP-c-Akt respectively. .. The cDNA encoding PTEN was excised with EcoRI from plasmid Pcmvpten [ ] [a gift from J.M.

Mutagenesis:

Article Title: A Type 2C Protein Phosphatase FgPtc3 Is Involved in Cell Wall Integrity, Lipid Metabolism, and Virulence in Fusarium graminearum
Article Snippet: .. The PCR product was digested with Hind III and Sac I, cloned into the pYES2 vector (Invitrogen), and transformed into the yeast mutant BY4741ΔPtc1. .. Yeast transformants were selected on synthetic medium lacking Ura (Clontech).

Article Title: The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea
Article Snippet: .. The full-length cDNA of BcATG8 was cloned into the yeast expression vector pYES2 (Invitrogen, Carlsbad, CA) and transformed into the yeast ATG8 null mutant using the lithium acetate (LiAc)/single-stranded DNA (ssDNA)/polyethylene glycol (PEG) transformation protocol ( ). .. Yeast transformants were selected on SD medium lacking uracil (Clontech, Palo Alta, CA).

Construct:

Article Title: Reconstitution of the mammalian PI3K/PTEN/Akt pathway in yeast
Article Snippet: .. Cassettes in which cDNAs encoding either c-Akt, c-AktK179M or an N-myristoylated c-Akt were fused in frame to an HA (haemagglutinin) epitope-tagged version of eGFP (enhanced green fluorescence protein) [HA–eGFP-Akt, HA–eGFP-AktK179M and myr-HA–eGFP-Akt respectively] were excised with HindIII and BamHI from the original Pcefl(X)-derived plasmids that were constructed for expression in mammalian cells [ ] (a gift from M. Lorenzo, Universidad Complutense, Madrid, Spain) and cloned into the corresponding sites in yeast vector pYES2 (Invitrogen), yielding plasmids pYES-GFP-c-Akt, pYES-GFP-c-AktK179M and pYES-myr-GFP-c-Akt respectively. .. The cDNA encoding PTEN was excised with EcoRI from plasmid Pcmvpten [ ] [a gift from J.M.

Polymerase Chain Reaction:

Article Title: A Type 2C Protein Phosphatase FgPtc3 Is Involved in Cell Wall Integrity, Lipid Metabolism, and Virulence in Fusarium graminearum
Article Snippet: .. The PCR product was digested with Hind III and Sac I, cloned into the pYES2 vector (Invitrogen), and transformed into the yeast mutant BY4741ΔPtc1. .. Yeast transformants were selected on synthetic medium lacking Ura (Clontech).

Expressing:

Article Title: Reconstitution of the mammalian PI3K/PTEN/Akt pathway in yeast
Article Snippet: .. Cassettes in which cDNAs encoding either c-Akt, c-AktK179M or an N-myristoylated c-Akt were fused in frame to an HA (haemagglutinin) epitope-tagged version of eGFP (enhanced green fluorescence protein) [HA–eGFP-Akt, HA–eGFP-AktK179M and myr-HA–eGFP-Akt respectively] were excised with HindIII and BamHI from the original Pcefl(X)-derived plasmids that were constructed for expression in mammalian cells [ ] (a gift from M. Lorenzo, Universidad Complutense, Madrid, Spain) and cloned into the corresponding sites in yeast vector pYES2 (Invitrogen), yielding plasmids pYES-GFP-c-Akt, pYES-GFP-c-AktK179M and pYES-myr-GFP-c-Akt respectively. .. The cDNA encoding PTEN was excised with EcoRI from plasmid Pcmvpten [ ] [a gift from J.M.

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Article Title: Cloning and Functional Characterization of a Vacuolar Na+/H+ Antiporter Gene from Mungbean (VrNHX1) and Its Ectopic Expression Enhanced Salt Tolerance in Arabidopsis thaliana
Article Snippet: .. The CDS of VrNHX1 was cloned into yeast expression vector pYES2.0 (Invitrogen, Carlsbad, CA, USA) with restriction sites of KpnI and BamHI. .. The yeast strains were transformed with pYES2.0 empty vector (labeled as AXTYES2.0 strain) or pYESVrNHX1 recombinant plasmid (labeled as AXTVrNHX1 strain) by Lithium acetate method and selected on SC ura− medium.

Article Title: Functional Desaturase Fads1 (Δ5) and Fads2 (Δ6) Orthologues Evolved before the Origin of Jawed Vertebrates
Article Snippet: .. After restriction digest, the insert was cloned into pYES2 yeast expression vector (Invitrogen), and sequenced. .. The constructs pYES2-ScaFads1 and pYES2-ScaFads2 were transformed into Saccharomyces cerevisiae competent cells (strain InvSc, Invitrogen).

Article Title: The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea
Article Snippet: .. The full-length cDNA of BcATG8 was cloned into the yeast expression vector pYES2 (Invitrogen, Carlsbad, CA) and transformed into the yeast ATG8 null mutant using the lithium acetate (LiAc)/single-stranded DNA (ssDNA)/polyethylene glycol (PEG) transformation protocol ( ). .. Yeast transformants were selected on SD medium lacking uracil (Clontech, Palo Alta, CA).

Article Title: Stress-induced TRBP phosphorylation enhances its interaction with PKR to regulate cellular survival
Article Snippet: .. Wild-type PKR was subcloned into the pYES2 yeast expression vector (Invitrogen) as previously described for galactose inducible PKR expression. .. The constructs were introduced into InvSc1 yeast cells (Invitrogen) using the Clontech Yeast Transformation Kit.

Article Title: Friedelin Synthase from Maytenus ilicifolia: Leucine 482 Plays an Essential Role in the Production of the Most Rearranged Pentacyclic Triterpene
Article Snippet: .. The full length Maytenus ilicifolia OSC ORFs were cloned into the yeast expression vector pYES2 (Invitrogen), which is under the transcriptional control of galactose (GAL1 promoter). .. The pYES2-Mi FRS and pYES2-Mi CAS1- plasmids were transformed into VZL 1303, and the resulting strains were grown in 0.5 L of synthetic complete medium without uracil (SC-U).

Sequencing:

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Transformation Assay:

Article Title: A Type 2C Protein Phosphatase FgPtc3 Is Involved in Cell Wall Integrity, Lipid Metabolism, and Virulence in Fusarium graminearum
Article Snippet: .. The PCR product was digested with Hind III and Sac I, cloned into the pYES2 vector (Invitrogen), and transformed into the yeast mutant BY4741ΔPtc1. .. Yeast transformants were selected on synthetic medium lacking Ura (Clontech).

Article Title: The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea
Article Snippet: .. The full-length cDNA of BcATG8 was cloned into the yeast expression vector pYES2 (Invitrogen, Carlsbad, CA) and transformed into the yeast ATG8 null mutant using the lithium acetate (LiAc)/single-stranded DNA (ssDNA)/polyethylene glycol (PEG) transformation protocol ( ). .. Yeast transformants were selected on SD medium lacking uracil (Clontech, Palo Alta, CA).

Plasmid Preparation:

Article Title: A Type 2C Protein Phosphatase FgPtc3 Is Involved in Cell Wall Integrity, Lipid Metabolism, and Virulence in Fusarium graminearum
Article Snippet: .. The PCR product was digested with Hind III and Sac I, cloned into the pYES2 vector (Invitrogen), and transformed into the yeast mutant BY4741ΔPtc1. .. Yeast transformants were selected on synthetic medium lacking Ura (Clontech).

Article Title: Reconstitution of the mammalian PI3K/PTEN/Akt pathway in yeast
Article Snippet: .. Cassettes in which cDNAs encoding either c-Akt, c-AktK179M or an N-myristoylated c-Akt were fused in frame to an HA (haemagglutinin) epitope-tagged version of eGFP (enhanced green fluorescence protein) [HA–eGFP-Akt, HA–eGFP-AktK179M and myr-HA–eGFP-Akt respectively] were excised with HindIII and BamHI from the original Pcefl(X)-derived plasmids that were constructed for expression in mammalian cells [ ] (a gift from M. Lorenzo, Universidad Complutense, Madrid, Spain) and cloned into the corresponding sites in yeast vector pYES2 (Invitrogen), yielding plasmids pYES-GFP-c-Akt, pYES-GFP-c-AktK179M and pYES-myr-GFP-c-Akt respectively. .. The cDNA encoding PTEN was excised with EcoRI from plasmid Pcmvpten [ ] [a gift from J.M.

Article Title: Semi-Selective Fatty Acyl Reductases from Four Heliothine Moths Influence the Specific Pheromone Composition
Article Snippet: .. Functional single substrate assay in yeast Each full–length FAR ORF was amplified using gene–specific primers ( ) and cloned in the pYES2.1 expression vector downstream of the GAL1 promoter according to the instructions given by the manufacturer (Invitrogen) before confirmation by sequencing with the vector specific primers Gal1 and V5. .. The four pgFAR constructs and the sole pYES2.1 plasmid were transformed into the InvSc1 strain of S. cerevisiae (Invitrogen) and grown on SC–U plates with 0.7% YNB (w/o aa, with ammonium sulphate), and a drop–out medium lacking uracil (ForMedium™ LTD, Norwich, England), and 2% glucose.

Article Title: Cloning and Functional Characterization of a Vacuolar Na+/H+ Antiporter Gene from Mungbean (VrNHX1) and Its Ectopic Expression Enhanced Salt Tolerance in Arabidopsis thaliana
Article Snippet: .. The CDS of VrNHX1 was cloned into yeast expression vector pYES2.0 (Invitrogen, Carlsbad, CA, USA) with restriction sites of KpnI and BamHI. .. The yeast strains were transformed with pYES2.0 empty vector (labeled as AXTYES2.0 strain) or pYESVrNHX1 recombinant plasmid (labeled as AXTVrNHX1 strain) by Lithium acetate method and selected on SC ura− medium.

Article Title: Functional Desaturase Fads1 (Δ5) and Fads2 (Δ6) Orthologues Evolved before the Origin of Jawed Vertebrates
Article Snippet: .. After restriction digest, the insert was cloned into pYES2 yeast expression vector (Invitrogen), and sequenced. .. The constructs pYES2-ScaFads1 and pYES2-ScaFads2 were transformed into Saccharomyces cerevisiae competent cells (strain InvSc, Invitrogen).

Article Title: The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea
Article Snippet: .. The full-length cDNA of BcATG8 was cloned into the yeast expression vector pYES2 (Invitrogen, Carlsbad, CA) and transformed into the yeast ATG8 null mutant using the lithium acetate (LiAc)/single-stranded DNA (ssDNA)/polyethylene glycol (PEG) transformation protocol ( ). .. Yeast transformants were selected on SD medium lacking uracil (Clontech, Palo Alta, CA).

Article Title: Stress-induced TRBP phosphorylation enhances its interaction with PKR to regulate cellular survival
Article Snippet: .. Wild-type PKR was subcloned into the pYES2 yeast expression vector (Invitrogen) as previously described for galactose inducible PKR expression. .. The constructs were introduced into InvSc1 yeast cells (Invitrogen) using the Clontech Yeast Transformation Kit.

Article Title: Friedelin Synthase from Maytenus ilicifolia: Leucine 482 Plays an Essential Role in the Production of the Most Rearranged Pentacyclic Triterpene
Article Snippet: .. The full length Maytenus ilicifolia OSC ORFs were cloned into the yeast expression vector pYES2 (Invitrogen), which is under the transcriptional control of galactose (GAL1 promoter). .. The pYES2-Mi FRS and pYES2-Mi CAS1- plasmids were transformed into VZL 1303, and the resulting strains were grown in 0.5 L of synthetic complete medium without uracil (SC-U).

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  • 94
    Thermo Fisher pyes2 ct vector
    GC/FID analysis of FAMEs isolated from Ole1 yeast cells expressing vFADs. After lyophilization the esterified fatty acids were transesterified with sodium methoxide and analyzed by GC/FID (see Materials and methods). ( a ) Chromatogram of the control yeast, Ole1 transformed with an empty <t>pYES2/CT</t> vector. ( b ) Chromatogram of the Ole1 yeast expressing vFAD-I (marked with a gray arrow in Figure 2 ). ( c ) Chromatogram of the Ole1 yeast expressing vFAD-II (marked with a black arrow in Figure 2 ). For the chromatogram of the InvSc2 strain (containing an active ole1 gene) see Supplementary Figure 5 .
    Pyes2 Ct Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pyes2 ct vector/product/Thermo Fisher
    Average 94 stars, based on 9 article reviews
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    pyes2 ct vector - by Bioz Stars, 2020-05
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    93
    Thermo Fisher pyes2 1 topo ta yeast expression kit
    S . cerevisiae growth under chloramphenicol selection. “Parental” refers to the parental, untransformed S . cerevisiae yeast strain 31019b. “Empty vector” refers to the parental yeast strain transformed with a <t>pYES2.1</t> vector carrying an eGFP coding region. “Chlo Resist” refers to the parental yeast strain transformed with a pYES2.1 vector carrying the chloramphenicol acyltransferase (CAT) resistance gene (ChloR). “+Chlo” indicates treatment of cultures with chloramphenicol. “+Ura” and “-URA” indicate the presence and absence of uracil supplement in the growth media respectively. Results are from five biological replicates. Asterisks indicate significantly different means with a p-value of
    Pyes2 1 Topo Ta Yeast Expression Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher resultant pyes2 crtiby
    Liquid chromatography–mass spectrometric analyses of carotenoids generated by yeasts expressing <t>crtIBY</t> from Aurantiochytrium sp. KH105. ( a ) Total ion chromatograms of carotenoids extracted from yeast cells carrying <t>pYES2</t> or pYES2- crtIBY and the β-carotene standard. An ion signal was selected within the molecular weight range of 537.44–537.45; ( b ) Mass spectrometry of carotenoids extracted from yeast cells carrying pYES2 or pYES2- crtIBY and the β-carotene standard at the retention times of 13.35, 13.21, and 13.22 min, respectively.
    Resultant Pyes2 Crtiby, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GC/FID analysis of FAMEs isolated from Ole1 yeast cells expressing vFADs. After lyophilization the esterified fatty acids were transesterified with sodium methoxide and analyzed by GC/FID (see Materials and methods). ( a ) Chromatogram of the control yeast, Ole1 transformed with an empty pYES2/CT vector. ( b ) Chromatogram of the Ole1 yeast expressing vFAD-I (marked with a gray arrow in Figure 2 ). ( c ) Chromatogram of the Ole1 yeast expressing vFAD-II (marked with a black arrow in Figure 2 ). For the chromatogram of the InvSc2 strain (containing an active ole1 gene) see Supplementary Figure 5 .

    Journal: The ISME Journal

    Article Title: Cyanophage-encoded lipid desaturases: oceanic distribution, diversity and function

    doi: 10.1038/ismej.2017.159

    Figure Lengend Snippet: GC/FID analysis of FAMEs isolated from Ole1 yeast cells expressing vFADs. After lyophilization the esterified fatty acids were transesterified with sodium methoxide and analyzed by GC/FID (see Materials and methods). ( a ) Chromatogram of the control yeast, Ole1 transformed with an empty pYES2/CT vector. ( b ) Chromatogram of the Ole1 yeast expressing vFAD-I (marked with a gray arrow in Figure 2 ). ( c ) Chromatogram of the Ole1 yeast expressing vFAD-II (marked with a black arrow in Figure 2 ). For the chromatogram of the InvSc2 strain (containing an active ole1 gene) see Supplementary Figure 5 .

    Article Snippet: DNA fragments, as gBlocks Gene Fragments (IDT), were cloned into the pYES2/CT vector (Thermo Fisher Scientific, Waltham, MA, USA) using Eco RI and Not I sites in frame so that the gene is fused to the vector’s His-tag at the N terminus of the protein, and sequenced to confirm their identity.

    Techniques: Isolation, Expressing, Transformation Assay, Plasmid Preparation

    S . cerevisiae growth under chloramphenicol selection. “Parental” refers to the parental, untransformed S . cerevisiae yeast strain 31019b. “Empty vector” refers to the parental yeast strain transformed with a pYES2.1 vector carrying an eGFP coding region. “Chlo Resist” refers to the parental yeast strain transformed with a pYES2.1 vector carrying the chloramphenicol acyltransferase (CAT) resistance gene (ChloR). “+Chlo” indicates treatment of cultures with chloramphenicol. “+Ura” and “-URA” indicate the presence and absence of uracil supplement in the growth media respectively. Results are from five biological replicates. Asterisks indicate significantly different means with a p-value of

    Journal: PLoS ONE

    Article Title: The genetic intractability of Symbiodinium microadriaticum to standard algal transformation methods

    doi: 10.1371/journal.pone.0211936

    Figure Lengend Snippet: S . cerevisiae growth under chloramphenicol selection. “Parental” refers to the parental, untransformed S . cerevisiae yeast strain 31019b. “Empty vector” refers to the parental yeast strain transformed with a pYES2.1 vector carrying an eGFP coding region. “Chlo Resist” refers to the parental yeast strain transformed with a pYES2.1 vector carrying the chloramphenicol acyltransferase (CAT) resistance gene (ChloR). “+Chlo” indicates treatment of cultures with chloramphenicol. “+Ura” and “-URA” indicate the presence and absence of uracil supplement in the growth media respectively. Results are from five biological replicates. Asterisks indicate significantly different means with a p-value of

    Article Snippet: Construction of transformation plasmids Yeast expression plasmids for the chloramphenicol resistance gene function test were synthesized using the pYES2.1 TOPO TA Yeast Expression Kit.

    Techniques: Selection, Plasmid Preparation, Transformation Assay

    Liquid chromatography–mass spectrometric analyses of carotenoids generated by yeasts expressing crtIBY from Aurantiochytrium sp. KH105. ( a ) Total ion chromatograms of carotenoids extracted from yeast cells carrying pYES2 or pYES2- crtIBY and the β-carotene standard. An ion signal was selected within the molecular weight range of 537.44–537.45; ( b ) Mass spectrometry of carotenoids extracted from yeast cells carrying pYES2 or pYES2- crtIBY and the β-carotene standard at the retention times of 13.35, 13.21, and 13.22 min, respectively.

    Journal: Genes

    Article Title: A Possible Trifunctional β-Carotene Synthase Gene Identified in the Draft Genome of Aurantiochytrium sp. Strain KH105

    doi: 10.3390/genes9040200

    Figure Lengend Snippet: Liquid chromatography–mass spectrometric analyses of carotenoids generated by yeasts expressing crtIBY from Aurantiochytrium sp. KH105. ( a ) Total ion chromatograms of carotenoids extracted from yeast cells carrying pYES2 or pYES2- crtIBY and the β-carotene standard. An ion signal was selected within the molecular weight range of 537.44–537.45; ( b ) Mass spectrometry of carotenoids extracted from yeast cells carrying pYES2 or pYES2- crtIBY and the β-carotene standard at the retention times of 13.35, 13.21, and 13.22 min, respectively.

    Article Snippet: Resultant pYES2-crtIBY and pYES2 plasmids were introduced individually to Saccharomyces cerevisiae INVSc1 (Thermo Fisher Scientific) using the polyethylene glycol/lithium acetate method.

    Techniques: Liquid Chromatography, Generated, Expressing, Molecular Weight, Mass Spectrometry