PLEASE EXPLAIN AND SHOW WORK You are ready to set up your reaction. You are prov

Question

PLEASE EXPLAIN AND SHOW WORK

You are ready to set up your reaction. You are provided with a stock of plasmid DNA that is at a concentration of 1 mg/ml. You have stock enzyme buffer that is 10X (meaning it is ten-times more concentrated than you need it to be in the actual reaction). You have deionized water (dH_2O) to use, as needed. As noted above, you want to cut 2 mu g DNA in a reaction volume of 50 mu l. To make sure you get complete cutting, you'll use an excess of enzyme so that you use 2.5 units per microgram of DNA (2.5U/mu g). The ApeKI enzyme is supplied at 5,000U/ml. How many mu l of DNA will you add to your reaction tube? How many mu l of 10X buffer will you add to your reaction tube? How many mu l of ApeKI will you add to your reaction tube? How many mu l of dH_2O will you add to your reaction tube? At what temperature will you incubate the reaction?

Explanation / Answer

Concentration of DNA is 1mg/ml which means 1µg/µl and we need to add 2µg DNA so the volume of DNA we will use is 2µl

For 50µl reaction, we need to use 5µl of 10X buffer

Given the concentration of enzyme is 5000U/ml which means 5U/µl. Also, we need to use 2.5U for per µg DNA so we will use 5U for the reaction. 5U means 1 µl.

We are using the enzyme procured from NEB then the reaction temperature will be 75 degree celcius

DNA 2µl Enzyme 1µl 10X buffer 5 µl dH20water to make volume 50 µl 42µl Total volume 50µl

Related Questions

Navigate

• Browse (All)
• Browse P
• Subjects

---

---

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.