The following antibodies were used in our studies: mouse monoclonal anti-Flag (1:10000, F3165, Sigma-Aldrich), mouse monoclonal anti-HA (1:10 000, H9658, Sigma-Aldrich), rabbit polyclonal anti-53BP1 (IF 1:200, sc-22760, Santa Cruz Biotechnology), rabbit polyclonal anti-MDC1 (1:1000, ab11171, Abcam), rabbit polyclonal anti-BRCA1 (IF 1:100, BS6423, Bioworld), polyclonal anti-γH2AX (IF 1:200, BS4760, Bioworld), rabbit polyclonal anti-Histone H3 (1:10 000, BE3015, EASYBIO), anti-β-Tubulin (1:10 000, BE3212-10, EASYBIO), rabbit polyclonal anti-ESCO2 (1:1000, A301-689A, Bethyl), and mouse monoclonal anti-acetyl-SMC3 K105/106, Clone 21A7 (ChIP 2μg, MABE1073, Millipore). The following reagents were used in our studies: KU55933 (ATM kinase inhibitor, S1092, Selleck Chemicals), NU7441 (DNA-PK inhibitor, S2638, Selleck Chemicals), VE821 (ATR inhibitor, S8007, Selleck Chemicals), DNase I (D5025, Sigma) and bleomycin sulfate (HY-17565, MedChemExpress).
Be3015
Manufactured by EASYBIO
The BE3015 is a high-precision laboratory balance designed for accurate weighing of samples. It features a digital display and a durable construction for reliable performance in a laboratory setting.
Automatically generated - may contain errors
Lab products found in correlation
Ab11171, by Abcam (1 mentions)
H9658, by Merck Group (1 mentions)
Ku55933, by Selleck Chemicals (1 mentions)
Ve 821, by Selleck Chemicals (1 mentions)
Sc 22760, by Santa Cruz Biotechnology (1 mentions)
D5025, by Merck Group (1 mentions)
Nu7441, by Selleck Chemicals (1 mentions)
F3165, by Merck Group (1 mentions)
Dnase 1, by Merck Group (1 mentions)
Ab19898, by Abcam (1 mentions)
Ab173289, by Abcam (1 mentions)
Ab181616, by Abcam (1 mentions)
Z033401 2, by Agilent Technologies (1 mentions)
Ac251, by Beyotime (1 mentions)
Ab9610, by Merck Group (1 mentions)
Palbociclib, by MedChemExpress (1 mentions)
Ab1220, by Abcam (1 mentions)
P1040, by Solarbio (1 mentions)
Be6706, by EASYBIO (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
3 protocols using be3015
1
Antibody and Reagent Usage for DNA Damage Studies
2
Lentiviral Knockdown of CDK4/6 in DIPG
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The shRNA-expressing Lentivirus system was described previously [25 (link)]. To generate pLKO plasmids containing short hairpins against CDK4 and CDK6, the pLKO vector was ligated with the annealed oligos. The sequences of shRNA oligo pairs are shown in Supplementary Table 2. Luciferase-GFP were cloned into pLEX based lentivirus vector, which was used to establish DIPG cell lines with luciferase-GFP [26 ].
The antibodies used in this study were anti-H3K27 M (ABE219, Millipore), anti-H3K27me3 (61,017, active motif), anti-H3K27Ac (ab191, Abcam), anti-H3 (BE3015, Easybio), anti-CDK4 (AC251, Beyotime), anti-CDK6 (AC256, Beyotime), anti-P-RB-S780 (ab173289, Abcam), anti-Ki67-PE conjugate antibody (12–5698-80, eBioscience), anti-CD133 (ab19898, abcam), anti-Nestin (Rat-401,DSHB), anti-GFAP (Z033401–2, DAKO), anti-Olig2 (AB9610, Milipore) and anti-RB (ab181616, Abcam). Secondary antibodies for western blot (Goat anti mouse/rabbit) were from Jackson. Secondary antibodies for flow cytometry (Goat anti mouse/rabbit) were from Huaxingbio. Enhanced chemiluminescence system (Tanon 5200) was used for detection. Palbociclib (PD0332991) were purchased from MedChemExpress.
The antibodies used in this study were anti-H3K27 M (ABE219, Millipore), anti-H3K27me3 (61,017, active motif), anti-H3K27Ac (ab191, Abcam), anti-H3 (BE3015, Easybio), anti-CDK4 (AC251, Beyotime), anti-CDK6 (AC256, Beyotime), anti-P-RB-S780 (ab173289, Abcam), anti-Ki67-PE conjugate antibody (12–5698-80, eBioscience), anti-CD133 (ab19898, abcam), anti-Nestin (Rat-401,DSHB), anti-GFAP (Z033401–2, DAKO), anti-Olig2 (AB9610, Milipore) and anti-RB (ab181616, Abcam). Secondary antibodies for western blot (Goat anti mouse/rabbit) were from Jackson. Secondary antibodies for flow cytometry (Goat anti mouse/rabbit) were from Huaxingbio. Enhanced chemiluminescence system (Tanon 5200) was used for detection. Palbociclib (PD0332991) were purchased from MedChemExpress.
3
SDS-PAGE and Western Blot Analysis of Histone Modifications
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Harvested leaves of kaku4-2 mutant and WT were ground in liquid nitrogen and the powder was extracted with 2× SDS-PAGE loading buffer (P1040; Solarbio) at 100 °C for 10 min. The solution was centrifuged for 10 min at 12,000 rpm. Extracted proteins were separated on 10% SDS-PAGE gels and transferred to polyvinylidene difluoride (PVDF) membrane (Millipore, 0.22μm). Membranes were blocked in blocking buffer [5% milk dissolved in 1× TBST (Tris Buffered Saline with Tween 20)] at room temperature (24 °C) for 1 h. Membranes were incubated for 1.5 h at room temperature with antibodies against H3K4me3 (07-473; Millipore), H3K27me3 (07-449; Millipore), H3K9me2 (ab1220; Abcam), actin (AC009; ABclonal), and H3 (BE3015; EASYBIO) in blocking buffer. After washing five times in TBST for 3 min each, membranes were incubated for 1 h at room temperature with Anti-Rabbit VHH HRP (KTSM1322; AlpalifeBio) and Goat Anti-Mouse IgG, HRP Conjugated (CW0102S; CWBIO) diluted 1/2,000. Membranes were washed five times in TBST and incubated for 1 min in electrochemiluminescence (ECL) buffer (BE6706; EASYBIO).
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