Navigation Links
An Animal-Free DNase I


Digestion of DNA to undetectable levels is important in many molecular biology applications. For instance, accurate quantitation of RNA targets by RT-PCR requires the removal of contaminating genomic DNA targets. In vitro transcribed RNA is also frequently treated with DNase to remove template DNA in both research and pharmaceutical applications. Ambion now supplies a recombinant DNase I that is prepared in a host with little to no RNase activity. The host is non-animal derived and the enzyme is purified through a process devoid of any animal byproducts.


Isolating RNase-free DNase I is a Challenge
DNase I has historically been prepared from bovine pancreas, one of the richest sources of RNase activity (~1 mg RNase A per gram of tissue). Therefore, it is often hard to obtain DNase I sufficiently free of RNase that it will not compromise RNA analysis experiments. Providing a high quality, RNase-free DNase I is very important to Ambion. This is borne out in a recent Biotechniques article (Matthews et al, 2002) that compared several different manufacturer's DNase I products (see sidebar at right).


A Pure Source of DNase I
Ambion employs stringent purification protocols and qualifies our bovine-derived DNase I at concentrations 5X or greater than that recommended for use. Now Ambion goes a step further by providing an alternative to bovine DNase I, recombinant DNase I (rDNase I), that further eliminates residual RNase and other copurified contaminants. rDNase is a highly purified product derived from cloned bovine DNase I. The rDNase I is prepared in a host that has RNase levels that are 1 x 107 fold lower than bovine pancreas.

Safety of biological products is also an emerging issue. Eliminating animal tissues and animal-derived components from the production process prevents potential transmission of pathogens. Materials derived from bovine sources are of special concern, due to known transmission of bovine spongiform encephalopathy (BSE) to humans via ingestion of cow products. Regulatory agencies are increasingly restrictive in this regard, prompting the pharmaceutical industry to remove high risk raw materials from manufacturing. Ambion's rDNase I now addresses these issues by providing a recombinant replacement for pancreatic bovine DNase I.


rDNase I Performance
Ambion's rDNase I meets or exceeds the performance of native bovine DNase I. It digests DNA to oligonucleotide-sized products or shorter. Yet its reaction kinetics are identical to that of native DNase I. Thus, rDNase I can be directly substituted for native DNase I with equivalent units.

The recommended reaction conditions are the same as Ambion's standard DNase I. Incubate 1-2 U of rDNase I per 1 g DNA for 30 min at 37C in a buffer consisting of 10 mM Tris-HCl, pH 7.5, 2.5 mM MgCl2, 0.5 mM CaCl2.


Quantitative Removal of Contaminating DNA
To demonstrate the efficiency of rDNase I in removing residual DNA, we measured genomic DNA contamination by real-time PCR before and after DNase I treatment. A total of 0.5 g of human genomic DNA was incubated in 50 l, and digested with 2 U of DNase I or rDNase I in a volume of 50 l for 30 min at 37C. One-tenth of this reaction volume (equivalent to 50 ng genomic DNA) was added to a 25 l real-time PCR reaction. The efficiency of DNA degradation was monitored by comparing cycle thresholds (Ct), with and without DNase digestion. As shown in Figure 1B, both DNase I and rDNase I shifted the real-time PCR signal by an average of 13.4 Ct's, or about 11,000-fold, to less than a single copy genome equivalent (~2 pg genomic DNA).

Figure 1. Real-time PCR of Genomic DNA Samples Treated with rDNase I vs. Bovine Pancreatic DNase I. (A) Standard curve generated with 10 fold serial dilutions of human genomic DNA ranging from 100 ng to 10 pg using an ABI 7900 real-time PCR machine. The target amplified was human GAPDH. (B) GAPDH was amplified from three human genomic DNA samples (0.5 g), either treated with DNase I (Ct=36.1), treated with rDNase I (Ct =36.9), or untreated (Ct= 23.1). The two DNase I treatments gave essentially identical results and were therefore averaged and compared to the untreated sample. The Ct of the averaged DNase I treatments vs. untreated human genomic DNA was 13.4.


Lack of RNase Activity
Since rDNase I is derived from a substantially "cleaner" source than pancreatic DNase I, the enzyme can be more readily purified to a RNase-free state. Although the recommended usage conditions for DNase I rarely require more than 2U per reaction, rDNase I passes Ambion's stringent RNase contamination quality control assay using 10 times as much enzyme for a 32-fold longer incubation! This result means that rDNase I is qualified RNase-free at 320 times typical usage conditions, creating an unsurpassed margin of safety for every application.


back to top


Ordering Information
For prices and availability, please contact our Customer Service Department.
Cat# Product Name Size 2235 Recombinant DNase I (RNase-free) 1000 U
'"/>

Source:


Page: All 1 2 3 4

Related biology technology :

1. The Worlds Best DNase Improved TURBO DNA-free
2. TURBO DNase: Hyperactive, Salt Tolerant DNase
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:10/11/2017)... and LAGUNA HILLS, Calif. , Oct. ... Cancer Research, London (ICR) and University ... SKY92, SkylineDx,s prognostic tool to risk-stratify patients with multiple myeloma ... MUK nine . The University of Leeds ... partly funded by Myeloma UK, and ICR will perform the ...
(Date:10/10/2017)... ... 2017 , ... San Diego-based team building and cooking events company, Lajollacooks4u, has ... The bold new look is part of a transformation to increase awareness, appeal to ... period. , It will also expand its service offering from its signature gourmet cooking ...
(Date:10/10/2017)... 10, 2017 International research firm Parks Associates announced ... at the TMA 2017 Annual Meeting , October 11 in ... residential home security market and how smart safety and security products impact ... Parks Associates: Smart Home ... "The residential security market has ...
(Date:10/9/2017)... DIEGO , Oct. 9, 2017  BioTech ... biological mechanism by which its ProCell stem cell ... critical limb ischemia.  The Company, demonstrated that treatment ... amount of limbs saved as compared to standard ... the molecule HGF resulted in reduction of therapeutic ...
Breaking Biology Technology:
(Date:3/28/2017)... , March 28, 2017 The ... Hardware (Camera, Monitors, Servers, Storage Devices), Software (Video Analytics, ... Region - Global Forecast to 2022", published by MarketsandMarkets, ... 2016 and is projected to reach USD 75.64 Billion ... and 2022. The base year considered for the study ...
(Date:3/24/2017)... 24, 2017 The Controller General of Immigration from ... Abdulla Algeen have received the prestigious international IAIR Award for the ... Continue Reading ... ... Controller Abdulla Algeen (small picture on the right) have received the IAIR ...
(Date:3/23/2017)... The report "Gesture Recognition and Touchless Sensing Market by Technology (Touch-based ... to 2022", published by MarketsandMarkets, the market is expected to be worth USD ... 2022. Continue Reading ... ... ...
Breaking Biology News(10 mins):