REPLI-g UltraFast Mini Kit
For ultrafast highly uniform whole genome amplification from small or precious samples
A lack of sufficient quantities of genomic DNA for genomic analysis can be overcome by global amplification of all DNA within a sample (whole genome amplification). The REPLI-g UltraFast Kit includes REPLI-g UltraFast DNA Polymerase, which enables highly uniform and accurate whole genome amplification from small samples in just 60–90 minutes, resulting in typical DNA yields of 7 μg per 20 μl reaction. A variety of starting materials can be used, including genomic DNA, fresh or dried blood, buccal swabs, fresh or frozen tissue, and cells. The DNA generated is suitable for direct use in a range of genetic analyses.
The average DNA yield is 7 µg per 20 µl reaction after 90 minutes with standard quality samples. If time is very limited, sufficient product is available for downstream genetic analysis after just 45 minutes (see figure "Consistent and high yields of amplified DNA"). Negative control samples (without template DNA) are not amplified, providing a reliable indicator of the presence/absence of sample or contaminating DNA (see figure "Consistent and high yields of amplified DNA").
Researcher feedback"In the past two years, we have been using the Repli-g Ultra Fast Mini Kit extensively for multiple projects. The multiple displacement amplification performed with the Repli-g Kit was able to faithfully amplify all DNA products at very low error rate and high coverage. The downstream applications we have used the kit for include most often NGS, but also qPCR and microarray. Moreover, we are working on implementing this technology in microfluidic or lab-on-a-chip systems, since the method requires no sophisticated thermal cycling program." Dr. Rong Fan, Assistant Professor of Biomedical Engineering, Yale University, US.
The availability of sufficient quantities of genomic DNA for genomic analysis is often lacking. Whole genome amplification overcomes this limitation by global amplification of all DNA within a sample, providing sufficient quantities to perform all analyses on the same DNA sample. REPLI-g UltraFast technology provides fast and highly uniform DNA amplification across the entire genome, with minimum sequence bias. The method is based on multiple displacement amplification (MDA) technology — isothermal genome amplification utilizing a new, uniquely processive DNA polymerase that gives DNA products of up to 100 kb in length in a much shorter time, while still avoiding the high sequence bias of PCR-based amplification methods. In addition, a unique alkaline denaturation buffer guarantees gentle denaturation of genomic DNA and avoids fragmentation of template DNA caused by heat denaturation methods. This ensures more sensitive and more reliable results in downstream assays.
Amplification of genomic DNA using the REPLI-g UltraFast Mini Kit involves 3 basic steps (see figure "Purified genomic DNA procedure"). First, the sample (10 ng purified genomic DNA, 0.5 µl whole blood, or 300 tissue culture cells) undergoes gentle alkaline denaturation, avoiding fragmentation and damage of template DNA. Next the sample is neutralized, and finally incubated with REPLI-g UltraFast DNA Polymerase at 30°C. The amplified DNA is ready to use after 60 minutes without further purification.
REPLI-g amplified genomic DNA can be used in a variety of downstream applications, including:
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