Journal: Integrative Cancer Therapies

Article Title: Analyses of BMAL1 and PER2 Oscillations in a Model of Breast Cancer Progression Reveal Changes With Malignancy

doi: 10.1177/1534735419836494

Figure Lengend Snippet: Endogenous mRNA expression of BMAL1 and PER2 in synchronized cell lines belonging to the MCF10 series. Relative mRNA expression levels were determined via RT-PCR, normalized to GAPDH , using 3 biological replicates with 3 technical replicates each. Error bars represent relative error.

Article Snippet: For generation of PER2- luciferase ( PER2:Luc ) plasmid, a 159-bp EcoRI/NotI fragment was isolated from a pGL3 basic PER2 construct, obtained from Addgene (plasmid #48747, deposited by Dr Joseph Takahashi).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction

Journal: Integrative Cancer Therapies

Article Title: Analyses of BMAL1 and PER2 Oscillations in a Model of Breast Cancer Progression Reveal Changes With Malignancy

doi: 10.1177/1534735419836494

Figure Lengend Snippet: Circadian oscillations of (A) BMAL1:Luc and (B) PER2:Luc in the MCF10 series of cells. Shown for each cell line are the 21 time series for BMAL1 and 20 for PER2 obtained, de-trended and de-noised using a discrete wavelet transform. Both reporters show stable oscillations across cellular transformations from nonmalignant MCF10A (blue) to premalignant MCF10AT.Cl2 (orange) to malignant MCF10Ca1h (yellow) and MCF10Ca1a cells (purple). However, BMAL1 has more time-series out of phase with the majority (gray; N = 4 for MCF10A, N = 2 for MCF10A.Cl2, N = 4 for MCF10Ca1h, and N = 6 for MCF10Ca1a) versus PER2 (gray; N = 5 for MCF10A).

Article Snippet: For generation of PER2- luciferase ( PER2:Luc ) plasmid, a 159-bp EcoRI/NotI fragment was isolated from a pGL3 basic PER2 construct, obtained from Addgene (plasmid #48747, deposited by Dr Joseph Takahashi).

Techniques:

Journal: Integrative Cancer Therapies

Article Title: Analyses of BMAL1 and PER2 Oscillations in a Model of Breast Cancer Progression Reveal Changes With Malignancy

doi: 10.1177/1534735419836494

Figure Lengend Snippet: Peaks and troughs of normalized (A) BMAL1:Luc and (B) PER2:Luc show that the first cycles of most oscillations are in phase and that PER2 and BMAL1 are antiphase. Heat maps are included, representing low (blue) to high (yellow) bioluminescence with one row per time series, organized by cell line (color bar on right indicating blue for MCF10A, orange for MCF10AT.Cl2, yellow for MCF10Ca1h, and purple for MCF10Ca1a cells). Black bars indicate recordings where no data were recorded at that time. Peaks (black triangles) and troughs (white triangles with black outline) are indicated for each recording and are used to determine outliers (for BMAL1:Luc , outliers trough after t = 48 hours, for PER2:Luc , outliers peak before t = 35 hours). Outliers are indicated by a gray entry in the first column, versus white for non-outliers.

Article Snippet: For generation of PER2- luciferase ( PER2:Luc ) plasmid, a 159-bp EcoRI/NotI fragment was isolated from a pGL3 basic PER2 construct, obtained from Addgene (plasmid #48747, deposited by Dr Joseph Takahashi).

Techniques:

Journal: Integrative Cancer Therapies

Article Title: Analyses of BMAL1 and PER2 Oscillations in a Model of Breast Cancer Progression Reveal Changes With Malignancy

doi: 10.1177/1534735419836494

Figure Lengend Snippet: Continuous wavelet transforms (CWT) estimate circadian periods for (A) BMAL1 and (B) PER2 over time. Shown are Morlet-wavelet estimates of the period of oscillation for each cell line and reporter (blue for MCF10A, orange for MCF10AT.Cl2, yellow for MCF10Ca1h, and purple for MCF10Ca1a). The periods of outlier time series are shown in gray.

Article Snippet: For generation of PER2- luciferase ( PER2:Luc ) plasmid, a 159-bp EcoRI/NotI fragment was isolated from a pGL3 basic PER2 construct, obtained from Addgene (plasmid #48747, deposited by Dr Joseph Takahashi).

Techniques: