DNase I (Rnase Free)(5u/ul)
Cat No: HC4007A
DNase I (Deoxyribonuclease I) is an endodeoxyribonuclease that can digest single- or double- stranded DNA. It recognizes and cleaves phosphodiester bonds to produce monodeoxynucleotides or single- or double- stranded oligodeoxynucleotides with phosphate groups at the 5'-terminal and hydroxyl at the 3'-terminal. The activity of DNase I depends on Ca2+ and can be activated by divalent metal ions such as Mn2+ and Zn2+. 5 mM Ca2+ protects the enzyme from hydrolysis. In the presence of Mg2+, the enzyme could randomly recognize and cleave any site on any strand of DNA. In the presence of Mn2+, the double strands of DNA can be simultaneously recognized and cleaved at almost the same site to form flat end DNA fragments or sticky end DNA fragments with 1-2 nucleotides protruding.
Components
|
Name |
0.1KU |
1KU |
5 KU |
50 KU |
|
DNase I, RNase-free |
20μL |
200μL |
1mL |
10 mL |
|
10×DNase I Buffer |
1mL |
1mL |
5 × 1mL |
5 ×10 mL |
Storage conditions
-25℃~-15℃ for storage; Transport under ice packs.
Instructions
1. Prepare the reaction solution in the RNase-free tube according to the proportions listed below:
|
Component |
Volume |
|
RNA |
X µg |
|
10 × DNase I Buffer |
1μL |
|
DNase I, RNase-free(5U/μL) |
1 U per µg RNA① |
|
ddH2O |
Up to 10μL |
Note: ①Calculate the volume of DNase I that needs to be added based on the amount of RNA.
2. 37 ℃ for 15 minutes;
3. Add 0.5M EDTA to the final concentration of 2.5mM~5mM, and heat at 65℃ for 10 minutes to stop the reaction. The sample can be directly used for the next reaction such as reverse transcription experiment.
Unit Definition
One unit is defined as the amount of enzyme which will completely degrade 1µg of pBR322 DNA in 10 minutes at 37℃.
Quality Control
RNase: 5U of DNase I with 1.6µg MS2 RNA for 4 hours at 37℃ yields no degradation as determined by agarose gel electrophoresis.
Notes
1. Please prepare 0.5MEDTA by yourself.
2. Use 1U DNase I per µg of RNA. However, if the RNA is less than 1µg, please use 1U DNase I.
3. Please place the enzyme on ice during operation.
