custom stellaris oligonucleotide sets  (Biosearch Technologies Inc)

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: (A) Cdc25 protein concentration doubles during G2. Cells expressing Cdc25-Rluc andAde4-Bluc from their endogenous loci (yFS874) were elutriation synchronized in early G2and followed through two synchronous cell cycles. Samples were taken every 20 minutesfor luciferase quantitation and examined microscopically for septation. As a comparison,cells expressing Wee1-Rluc and Ade4-Bluc (yFS810) were similarly assayed. Themidpoint of septation for each cycle is marked with an arrowhead and the inferred M-G1-S phases of the cycles are indicated in gray. The mean and standard error of the Ade4-nomalized Cdc25 and Wee1 signal, relative to time 0. n = 3. See also Figures S1 and S3.

Article Snippet: Cells were permeabilized in 70% EtOH, pre-blocked in BSA and salmon sperm DNA, and incubated over-night with custom Stellaris oligonucleotides sets (Biosearch Technologies) designed against cdc25 (CAL Fluor Red 610) and rpbl (Quasar 670) mRNAs ( Table S2 ).

Techniques: Protein Concentration, Expressing, Luciferase, Quantitation Assay, Comparison

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: (A) cdc25 transcript concentration doubles during G2. Wild-type cell (yFS105) wereelutriation synchronized in early G2 and followed through two synchronous cell cycles.Samples were taken every 20 minutes for RNA quantitation and examinedmicroscopically for septation. Steady-state mRNA levels were determined using theNanoString nCounter method with custom probes and normalized to total mRNA counts.Data points represent independent biological replicates, the lines connect the mean ofthose points. The midpoint of septation for each cycle is marked with an arrowhead andthe inferred M-G1-S phases of the cycles are indicated in gray. For the first two hours,n=2, for the rest of the time course, n=1.

Article Snippet: Cells were permeabilized in 70% EtOH, pre-blocked in BSA and salmon sperm DNA, and incubated over-night with custom Stellaris oligonucleotides sets (Biosearch Technologies) designed against cdc25 (CAL Fluor Red 610) and rpbl (Quasar 670) mRNAs ( Table S2 ).

Techniques: Concentration Assay, Quantitation Assay

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: (A) The delay in Cdc25 expression after a cycloheximide pulse mirrors the delay inmitotic entry. Cells expressing Cdc25-Bluc and Wee1-Rluc (yFS870) were elutriationsynchronized in early G2. Samples were taken every 20 minutes for luciferasequantitation and examined microscopically for septation. At the indicated times beforedivision of the untreated cells, the culture was split and one half was treated with 100μg/ml cycloheximide for 20 minutes.

Article Snippet: Cells were permeabilized in 70% EtOH, pre-blocked in BSA and salmon sperm DNA, and incubated over-night with custom Stellaris oligonucleotides sets (Biosearch Technologies) designed against cdc25 (CAL Fluor Red 610) and rpbl (Quasar 670) mRNAs ( Table S2 ).

Techniques: Expressing

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: (A) A Cdc25-concentration-regulated model of the cell cycle. Simulation of the cell cycle using an ordinary differential equation model in which the size-dependent expression of Cdc25 triggers entry into mitosis at the appropriate size.

Article Snippet: Cells were permeabilized in 70% EtOH, pre-blocked in BSA and salmon sperm DNA, and incubated over-night with custom Stellaris oligonucleotides sets (Biosearch Technologies) designed against cdc25 (CAL Fluor Red 610) and rpbl (Quasar 670) mRNAs ( Table S2 ).

Techniques: Concentration Assay, Expressing

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: Key Resources Table

Article Snippet: Cells were permeabilized in 70% EtOH, pre-blocked in BSA and salmon sperm DNA, and incubated over-night with custom Stellaris oligonucleotides sets (Biosearch Technologies) designed against cdc25 (CAL Fluor Red 610) and rpbl (Quasar 670) mRNAs ( Table S2 ).

Techniques: Recombinant, Reporter Assay, Quantitative RT-PCR, Software

Journal: Current biology : CB

Article Title: Size-Dependent Expression of the Mitotic Activator Cdc25 Suggests a Mechanism of Size Control in Fission Yeast

doi: 10.1016/j.cub.2017.04.016

Figure Lengend Snippet: Key Resources Table

Article Snippet: Stellaris oligonucleotides sets , Biosearch Technologies , N/A.

Techniques: Recombinant, Reporter Assay, Quantitative RT-PCR, Software