Sybr gold

Manufactured by Bio-Techne

SYBR Gold is a highly sensitive fluorescent nucleic acid stain used for the detection and quantification of DNA in various applications, such as gel electrophoresis and real-time PCR. It exhibits high sensitivity and can detect as little as 125 pg of double-stranded DNA.

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Lab products found in correlation

Trevigen comet assay kit, by Bio-Techne (3 mentions) Eclipse 80i, by Nikon (1 mentions) Comet assay kit, by Bio-Techne (1 mentions)

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SYBR gold solution (2 citations)

4 protocols using sybr gold

DNA Damage Quantification by Comet Assay

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The neutral and alkaline comet assay were performed on the Trevigen Comet Assay kit (Cat. 4252-040-K, Trevigen, Gaithersburg, Maryland, USA). Cells were pretreated with 0.5 mM VPA for 24 h followed by 8 Gy IR treatment. Low melting agar was combined with cell suspension at a ratio of 1:10 after treatments, then spread onto comet slide immediately. The slides were cooled for 30 min, then immersed in lysis solution. Slides were immersed into cold alkaline unwinding solution for 1 h (alkaline comet), then transferred into neutral or alkaline electrophoresis buffer at 21 V for 30 min at 4°C in the dark. The slides were washed twice in distilled water and 70% ethanol for 5 min, then dried and stained with SYBR gold (1:10000, Trevigen).

Liu G., Lim D., Cai Z., Ding W., Tian Z., Dong C., Zhang F., Guo G., Wang X., Zhou P, & Feng Z. (2021). The Valproate Mediates Radio-Bidirectional Regulation Through RFWD3-Dependent Ubiquitination on Rad51. Frontiers in Oncology, 11, 646256.

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Neutral Comet Assay of Fallopian Epithelial Cells

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The neutral comet assay was performed using the Trevigen Comet Assay kit (Trevigen, Gaithersburg, MD). Fallopian epithelial cells (FT33-TAg) were grown in the presence of individual FF samples, matched patient plasma, USG, no USG, or carboplatin (1 μM). After 24 hr incubation, the cells were suspended in cold PBS, and an aliquot (1000 cells/10 μl) was added to 100 μl of LMA agarose maintained at 39 °C and spread onto a comet slide. The slide was incubated at 4 °C for 10 min and transferred to cold lysis solution for 60 min at 4 °C. A denaturation step was performed in 50 mM Tris base, 150 mM pH 9, for 30 min at 4 °C. The slides were then subjected to electrophoresis with cold TAE buffer, pH 8.2 at 25 V for 30 min at 4 °C, and immersed in DNA precipitation solution (100 mM NH4Ac in 95% ethanol) and then in 100% ethanol for 30 min and air dried. DNA was stained with 100 μl SYBR Gold (Trevigen, 1:30,000) for 20 min and immediately rinsed with dH2O and air dried. The slides were imaged using an upright Nikon Eclipse 80i fluorescence microscope at ×20, and analyzed using Casplab software (Casplab.com) [25 (link)].

Brachova P., Alvarez N.S., Van Voorhis B.J, & Christenson L.K. (2017). Cytidine deaminase Apobec3a induction in fallopian epithelium after exposure to follicular fluid. Gynecologic oncology, 145(3), 577-583.

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Comet Assay for DNA Damage Analysis

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The neutral comet assay was performed using the Trevigen Comet Assay kit (Trevigen, Gaithersburg, MD). At the indicated time-points after irradiation, the cells were suspended in cold PBS, and an aliquot of cells (10³/10 μl) was added to 100 μl of molten LMA agarose maintained at 39°C. An aliquot of 10 μl was immediately spread onto each comet slide at 37°C. The slide was incubated at 4°C for 10 min to accelerate gelling of the agarose and then transferred to cold lysis solution for 60 min at 4°C. A denaturation step was performed in 50 mM Tris base, 150 mM CH₃COONa₃H₂O, pH 9, for 30 min at 4°C. The slides were then subjected to electrophoresis with cold TAE buffer, pH 8.2 at 25 V for 30 min at 4°C, and immersed in DNA precipitation solution (100 mM NH₄Ac in 95% ethanol) and then in 100% ethanol for 30 min and air dried. DNA was stained with 100 μl SYBR Gold (Trevigen, 1:30,000) for 20 min and immediately rinsed with dH₂O and air dried. The slides were analyzed using an All-in-One BZ 9000 fluorescence microscope and CometScore free ware v1.5 (TriTek, Niigata, Japan).

Kurashige T., Shimamura M, & Nagayama Y. (2016). Differences in quantification of DNA double-strand breaks assessed by 53BP1/γH2AX focus formation assays and the comet assay in mammalian cells treated with irradiation and N-acetyl-L-cysteine. Journal of Radiation Research, 57(3), 312-317.

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Comet Assay for DNA Damage Evaluation

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Alkaline comet assay was performed using the CometAssay^® Kit (Trevigen). Before being subjected to irinotecan, the Caco2 and HT29 cell lines were first transfected. The cells were then suspended in cold PBS and mixed with molten LMAgarose (37°C preheat) at a ratio of 1:10 and quickly pipetted 50 μl was quickly pipetted onto CometSlide™, ensuring that the sample completely covered the sample area. The slides were incubated at 4°C for about 20 min to allow the agarose to gel before being immersed in 4°C Lysis Solution for 1 h. The slides were then transferred into Alkaline Unwinding Solution and incubated at room temperature for 20 min. The slides were subjected to electrophoresed at 21V for 40 min and gently immersed twice in dH₂O and once in 70% ethanol for 5 min each. Next, the samples were air‐dried and 50 μl diluted SYBR^® Gold (Trevigen) was added to every sample and stained in dark place at room temperature for 30 min. The slides were then gently rinsed in water, air‐dried and observed using a fluorescent microscope. CaspLab Software was used to analyse the length of the DNA tails.

Mei C., Sun Z., Tan L., Gong J., Li X, & Liu Z. (2022). eIF3a‐PPP2R5A‐mediated ATM/ATR dephosphorylation is essential for irinotecan‐induced DNA damage response. Cell Proliferation, 55(4), e13208.

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