Concentration of purified DNA was measured using a spectrophotometer (NanoDrop 1000, Thermo Fisher Scientific). For ligations with T4 DNA ligase (Quick Ligation Kit, NEB), 50–100 ng backbone DNA (DNA fragment containing the DNA replication origin) was used in 20-µl volume, with molar 1:1–3 backbone:insert ratios, at room temperature for 5–10 min. Gibson assemblies were performed with ~75 ng backbone DNA and a molar 1:1–5 backbone:insert ratios in a 15-µl reaction volume, using NEBuilder HiFi DNA Assembly Master Mix (2×) (NEB) for 15–60 min at 50 °C.
Nebuilder hifi dna assembly master mix 2
Manufactured by New England Biolabs
NEBuilder HiFi DNA Assembly Master Mix (2×) is a high-fidelity, two-component DNA assembly solution. It enables the seamless joining of multiple DNA fragments in a single reaction.
Automatically generated - may contain errors
3 protocols using nebuilder hifi dna assembly master mix 2
1
DNA Quantification and Ligation Optimization
2
Plasmid DNA Ligation and Assembly
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Concentrations of DNA fragments were measured with a spectrophotometer (NanoDrop 1000; Thermo Fisher Scientific). Ligations were performed with 30–100 ng plasmid backbone DNA (an ori-containing DNA fragment) in a 20 µl reaction volume at room temperature, with 1:1–3 backbone:insert molar ratios, using T4 DNA Ligase (Quick Ligation Kit; NEB) for 5–10 min. Isothermal Gibson assemblies were performed in a 15 µl reaction volume with 1:1–1:5 backbone:insert molar ratios and 50–75 ng backbone DNA, using NEBuilder HiFi DNA Assembly Master Mix (2×; NEB) for 20–60 min at 50 °C.
3
DNA Ligation and Gibson Assembly
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Agarose-gel purified DNA fragment concentrations were determined using a spectrophotometer (NanoDrop 1000, Thermo Fisher Scientific). Ligations were performed using 50–100 ng backbone DNA (DNA fragment containing the origin of replication) in a volume of 20 µl, with molar 1:1–3 backbone:insert ratios, using T4 DNA ligase (Quick Ligation Kit, NEB) at room temperature for 5–10 min. Gibson assemblies were performed using 75 ng backbone DNA in a 15 µl reaction volume and a molar 1:1–5 backbone:insert ratios, using the NEBuilder HiFi DNA Assembly Master Mix (2×) (NEB) for 20–60 min at 50 °C.
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!