The total volume of extracted DNA was precipitated with 1/10th volume of 3M sodium acetate (Invitrogen, Carlsbad, California) and a double volume of chilled molecular-biology-grade ethanol (Sigma-Aldrich, St. Louis, Missouri) at -20°C overnight. The precipitated DNA was pelleted down at 15,000 rpm for 10 min at 4°C. The supernatant was discarded, and the pellet was tapped, dislodged, and washed twice with 700 μL of 70% ethanol, followed by centrifugation at 15,000 rpm for 10 min at 4°C. The clean DNA pellet was dried in a sterile environment at 37°C for 1 h or until dry. The DNA pellet was dissolved in 3.125 μL/12.5 μL of DNA suspension buffer (Teknova, Hollister, California) as needed. The re-suspended DNA was stored at -20°C until further use.
Molecular biology grade ethanol
Manufactured by Merck Group
Molecular-biology-grade ethanol is a high-purity ethanol solution specifically designed for use in molecular biology applications. It is a clear, colorless liquid with a characteristic odor. This product meets the stringent quality standards required for sensitive molecular techniques, ensuring consistent performance and reliable results.
Automatically generated - may contain errors
Lab products found in correlation
Dna suspension buffer, by Teknova (2 mentions)
Poly a carrier dna, by Roche (2 mentions)
Qiaamp dna mini kit, by Qiagen (2 mentions)
Thermomixer, by Eppendorf (2 mentions)
Qiaamp dna blood mini kit, by Qiagen (2 mentions)
Nanodrop, by Thermo Fisher Scientific (2 mentions)
20 hydroxyeicosatetraenoic acid 20 hete, by Cayman Chemical (1 mentions)
4t1 murine breast cancer cell, by American Type Culture Collection (1 mentions)
8 protocols using molecular biology grade ethanol
1
DNA Precipitation and Purification Protocol
2
Oligonucleotide Purification and Storage
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All oligonucleotides were synthesized by the Stanford Protein and Nucleic Acids facility (PAN) using standard phosphoramidite chemistry. Oligonucleotides were purified by denaturing 7M urea polyacrylamide gel electrophoresis (PAGE) in 1× TBE, followed by overnight elution from an excised band at 4°C in 300 mM Sodium Acetate pH 5.2 and 1 mM EDTA. DNA was precipitated and recovered by adding 100% molecular biology grade ethanol (Sigma Aldrich) to 70% final v/v and centrifuged for 30 min at 14,000 × g and 4°C. Alternately, RNA-containing oligonucleotides were recovered by precipitation in 75% ethanol v/v and centrifuged at 14,000 × g for 30 min at 4°C. The ethanol mixture was aspirated and oligonucleotides were then washed with an equal volume of 70 or 75% ethanol and pelleted again for 10 min at 14,000 × g and 4°C. Ethanol was aspirated and the pellets were allowed to dry under vacuum to remove residual ethanol. Oligonucleotides were then resuspended in nuclease free water, quantified by Nanodrop (Thermo Scientific), and stored at −80°C.
3
Ethanol Exposure in Differentiated Podocytes
4
Bacterial DNA Extraction from Various Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA extraction was performed from 1 mL of the different sample types using a QIAamp DNA mini kit (Qiagen, Hilden, Germany) as per the vendor’s instructions with slight modifications. All samples were pelleted down at 15,000 rpm for 10 min at 4°C, and the supernatant was discarded. The pellets underwent bacterial DNA isolation via resuspension in 180 μL of Buffer ATL and 20 μL of proteinase K. The lysate was incubated for 1 h at 56°C in a thermomixer (Eppendorf, Hamburg, Germany) with shaking. Subsequently, 200 μL of Buffer AL and 10 μg of poly(A) carrier DNA (Roche, Basel, Switzerland) were added to the lysate, which was incubated for an additional 10 min at 70°C. Next, 230 μL of molecular-biology-grade ethanol (Sigma-Aldrich, St. Louis, Missouri) was added to the lysate, which was then passed through a QIAamp mini spin column at 6000 g for 1 min at RT. The filtrate was discarded, and the spin column was washed with wash buffers AW1 and AW2, as per the vendor’s instructions. Finally, DNA was eluted twice with 150 μL of Buffer AE (pre-heated to 65°C), resulting in a total volume of ~300 μL. Bacterial DNA extraction from WB was conducted using a QIAamp DNA blood mini kit (Qiagen, Hilden, Germany) following the vendor’s recommended protocols.
5
Ethanol Precipitation of DNA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total volume of extracted DNA was precipitated with 1/10 th volume of 3M sodium acetate (Invitrogen, Carlsbad, California) and double volume of chilled molecular biology grade ethanol (Sigma-Aldrich, St. Louis, Missouri) at -20°C overnight. The precipitated DNA was pelleted down at 15,000rpm for 10 minutes at 4°C. The supernatant was discarded and the pellet washed twice with 700 µl of 70% ethanol by tapping and dislodging the pellet followed by centrifugation at 15,000rpm for 10 minutes at 4°C. The clean DNA pellet was dried in a sterile environment at 37°C for 1 hour or until dry. The DNA pellet was dissolved in 3.125µl/12.5µl of DNA suspension buffer (Teknova, Hollister, California) according to need. The re-suspended DNA was stored at -20°C until further use.
6
Bacterial DNA Extraction from Diverse Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA extraction was done from 1 ml of the different sample types using QIAamp DNA mini kit (Qiagen, Hilden, Germany), as per the vendor's instructions with slight modifications. All the samples were pelleted down at 15,000rpm for 10 minutes at 4°C and the supernatant was discarded. The pellets underwent bacterial DNA isolation by re-suspending in 180µl of Buffer ATL and 20µl of Proteinase K. The lysate underwent 1 hour of incubation at 56°C in a thermomixer (Eppendorf, Hamburg, Germany) with shaking. Following this, 200µl of Buffer AL was added along with 10µg of poly (A) carrier DNA (Roche, Basel, Switzerland) to the lysate and incubated for an additional 10 minutes at 70°C. Then 230µl of molecular-biology grade ethanol (Sigma-Aldrich, St. Louis, Missouri) was added to the lysate before passing through a QIAamp mini spin column at 6000g for 1 minute at RT. The filtrate was discarded and the spin column was washed with wash buffers AW1 and AW2, as per the vendor's instructions. Finally DNA was eluted with 150µl of Buffer AE (pre-heated to 65°C) twice resulting in a total volume of ~300µl. Bacterial DNA extraction from whole blood was done with QIAamp DNA blood mini kit (Qiagen, Hilden, Germany) following vendor recommended protocols.
7
Murine Breast Cancer Cell Culture
8
FFPE Tumor Macro-dissection and STRAT4 Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
FFPE samples were processed according to the package insert instructions of the STRAT4 assay kit. For each specimen, one unstained slide was overlaid onto the H&E slide which had been marked by a pathologist to select the invasive carcinoma, and then was used to choose the material to be macro-dissected into a 1.5 mL eppendorf tube using a razor blade. Macro-dissected tumor material was then mixed with 1.2 mL of FFPE lysis reagent and 20 µL of proteinase K. The tubes containing the sample lysate were placed in heat blocks for incubation at 80 °C for 30 min. Sample lysate was then mixed with 1.2 mL of ethanol (molecular biology grade, Sigma-Aldrich). For each sample, 520 µL of the lysate was transferred to the sample chamber of a STRAT4 cartridge and placed into a GeneXpert module for RNA extraction, purification, and RT-qPCR analysis.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!