Ambion, Inc

The TURBO™ DNase Advantage

Table 1. TURBO DNase Activity* in Various Molecular Biology Buffers	% Activity in Common Buffers
	Wild-type DNase I	TURBO DNase
DNase Reaction Buffer	100	100
RT Buffer	0.6	23 (38)**
NEB Buffer #3	0.1	58 (580)
NEB Buffer #4	1.4	205 (146)
PBS (+5 mM MgCl₂)	0.005	16.7 (>100)

*DNase activity was measured using Ambion’s DNaseAlert™ QC Kit which uses an oligonucleotide substrate in a fluorometric assay.

**Values in parenthesis show the fold improvement in activity over wild-type bovine DNase I. Note that % activity levels <1% are difficult to measure with great accuracy.

A) TURBO DNase

B) Wild-type DNase I

Figure 1. TURBO DNase has 400% greater catalytic efficiency than wild-type DNase I. TURBO DNase (panel A) has a 6-fold lower K_m and a 400% greater catalytic efficiency (V_max/K_m) than the wild-type enzyme (panel B). Since the goal of every DNase I digestion is to reduce the substrate DNA concentration to essentially zero, the much lower K_m of TURBO DNase for DNA means that the mutant enzyme will always be more efficient than wild-type DNase in producing RNA samples that are free of contaminating DNA.

Salt Tolerance of TURBO DNase As a result of tighter DNA binding, TURBO DNase manifests a remarkable tolerance for monovalent salt. In fact, TURBO DNase is hyperactive in physiological salt concentrations (~150 mM), whereas wild-type DNase I is strongly inhibited. As shown in the following figure, the activity (V_max/K_m) of the TURBO DNase is increased 333% when 100 mM NaCl is added to 1X DNase I buffer. In contrast, the wild-type DNase activity is decreased 25-fold under the same conditions.


Figure 2. Hyperactivity of TURBO DNase in 0.1 M NaCl

	% of Peak Activity		Ratio
mM NaCl	wild-type DNase I	TURBO DNase	TURBO/ wild-type
0	100	39	0.4
10	72	47	0.7
20	51	56	1.1
50	22	77	3.5
75	9	94	10.4
100	4	100	25.0
125	2	99	49.5
150	1	82	82.0
200	1	44	44.0
400	0	2	N/A

Table 2. Activity of Wild-type vs. TURBO DNase I as a Function of Salt. Assay conditions: 37°C, 200 nM DNaseAlert™, 25 pM DNase I, 1X DNase I Buffer + NaCl as given.

DNase	Vendor	Relative Activity
TURBO DNase	Ambion	333
RQ DNase I	Promega	100
DNase I, Amplification Grade	Invitrogen	100

Table 3. TURBO DNase is More Potent than Competitor’s Wild-type DNase I Enzymes. Activity was measured using the DNaseAlert assay. Each enzyme was diluted to 0.008 U in the supplied 1X DNase I reaction buffer, and cleavage of the DNaseAlert substrate (200 nM) was monitored using a SpectraMAX GeminiXS Fluorometer (535/556 nm ex/em) after the addition of 5 µl of the diluted enzyme to a 95 µl reaction.