A More Uniform Spectrum

The new GeneMorph^® II random mutagenesis kit* overcomes the problem of mutational bias that can skew representation of random mutant libraries. Enzymes commonly employed in errorprone PCR include Taq, which is four times more likely to mutate As and Ts under error-prone conditions, and Mutazyme^® DNA polymerase, which is nearly three times more likely to mutate Gs and Cs than As and Ts. GeneMorph II random mutagenesis kit features the improved Mutazyme^® II DNA polymerase,** which mutates Gs and Cs at equivalent rates to As and Ts. In the Mutazyme II polymerase blend, Mutazyme polymerase and a novel Taq mutant, which exhibits a higher error rate compared to wild-type Taq, have been combined at a ratio that introduces all possible mutation types. As a result of reduced mutational bias, libraries created with GeneMorph II generally exhibit greater representation compared to libraries generated with other error-prone PCR enzymes (Figure 1).

Easily Control Mutation Frequency

Precise control over mutation frequency improves the likelihood of obtaining a mutant with desired properties. With Taq-based, error-prone PCR methods, this requires painstaking adjustments to PCR reaction conditions. In the GeneMorph II kit, low, medium, and high mutation frequencies are achieved using a single set of optimal reaction conditions (e.g., constant MgCl2 and balanced dNTPs). The only parameter that is varied is DNA template concentration, so the same spectrum of mutations is produced over a broad range of mutation frequencies (0.1 to 1.6% per PCR) (Figure 2).

Robust Yields Simplifies Library Construction

Error-prone PCR using Taq DNA polymerase is limited to 2 to 3 kb, depending on the target complexity. With Mutazyme II DNA polymerase, you can amplify a wide range of amplicon sizes up to 6 kb with robust yields, making cloning and library construction more efficient and successful (Figure 3).

Discover More, Faster

The GeneMorph II kit offers a fast and easy method for creating diverse mutant collections, with reduced bias compared to error-prone PCR methods employing Taq DNA polymerase. Additionally, you have complete control over mutation frequency without extra work. By screening more representative libraries, you can more rapidly uncover key sites responsible for structurefunction relationships and accomplish your research goals faster than ever before.

Figure 2
Simply Change the Input Amount for the Desired Frequency
Controlling the mutation frequency using the GeneMorph^® II kit is easy. Stratagene generated a range of mutation frequencies by adjusting the initial template amount from 16 pg to 1000 ng. By increasing the amount of template (in parenthesis) added to the PCR, the mutation frequency decreased from 16 mutations per kb using 16 pg to 3.8 mutations per kb using 1000 ng.

Figure 3
Mutazyme II DNA Polymerase Amplifies a Wide Range of Target Sizes
When compared to Taq DNA polymerase (under error-prone PCR conditions: 0.5mM Mn2+, 0.2mM dATP and dGTP, and 1mM dCTP and TTP), Mutazyme II amplifies a wider range of target sizes. In this comparison, Mutazyme II DNA polymerase amplified PCR products up to 4 kb, with more robust yield compared to Taq, and can be used reliably for amplicons up to 6 kb (data not shown). Lanes 1-6 are as follows: 1. Lac Z fragment (650 bp), 2. 1 kb pBSII clone target, 3. Pfu gene (2.3 kb), 4. Taq gene (2.6 kb), 5. 3.9 kb pBSII clone fragment, 6. 4 kb Pfu/pBSII fragment

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