FAQ-702
Are Centromeres and Telomeres amplified using REPLI-g WGA?
These regions are not amplified because only a subset of the random primers in the REPLI-g amplification mix can prime within these extensively repeated regions. Despite the high processivity of the Phi29 DNA Polymerase, centromeres and telomeres are at a competitive disadvantage during amplification, and drop out. In experiments we have done, single copy sequences within approximately 5000 bases of a poorly amplified region can be affected during amplification. If your gene is further than 5000 bases apart from a centromere or telomere, it should be amplified just fine.
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REPLI-g Single Cell Kit
DNA sequence analysis and genotyping of biological samples using innovative instrumentation, such as next-generation sequencing (NGS) platforms, is often limited by the small amount of sample available. The REPLI-g Single Cell Kit is specially designed to uniformly amplify genomic DNA from single cells (1 to <1000 bacterial or tumor cells) or purified genomic DNA with complete genome coverage. Additional protocols are also available for use with fresh or dried blood or fresh or frozen tissue. Dedicated buffers and reagents undergo a unique, controlled decontamination procedure to avoid amplification of contaminating DNA, ensuring highly reliable results every time. Accurate amplification of genomes with negligible sequence bias and no genomic drop-outs is achieved with innovative Multiple Displacement Amplification (MDA) technology. In contrast to PCR-based WGA technologies, high fidelity rates are increased up to 1000-fold, avoiding costly false positive or negative results.
REPLI-G Kits
Limitations on the number and type of genomic analyses that can be performed due to insufficient quantities of genomic DNA can be overcome by global amplification of all DNA within a sample (whole genome amplification). REPLI‑g Kits provide DNA polymerase, buffers and reagents for highly uniform whole genome amplification with minimal sequence bias and can be used with various starting materials, including genomic DNA, fresh or dried blood, buccal swabs, fresh or frozen tissue and cells. QIAGEN REPLI-g Kits are available in several different sizes and configurations which allows researchers to build the best workflow for their application and lab space. Typical REPLI-g yields range from approx. 7 µg to 40 µg of amplified DNA, depending on which kit and workflow is used. The REPLI-g Mini and Midi Kits offer a single tube workflow with higher yields of amplified DNA, while the REPLI-g UltraFast Kit enables highly uniform and accurate whole genome amplification in just 60–90 minutes. For high-throughput processing in 96-well format, the streamlined, room-temperature procedure of the REPLI-g Screening Kit enables straightforward liquid handling that requires just 20 minutes. The REPLI-g Screening kit is intended for rapid screening using a range of genotyping, PCR, sequencing or microarray assays.
REPLI-g FFPE Kit
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 FFPE Kit consists of DNA Polymerase, novel buffers, and reagents that enable efficient whole genome amplification from formalin-fixed and paraffin embedded (FFPE) tissue without the requirement for prior DNA purification. After lysis of the tissue section, the DNA is processed so that fragmented DNA is ligated. The long DNA strands created are amplified using proven REPLI-g technology.
REPLI-g Mitochondrial DNA Kit
The REPLI-g Mitochondrial DNA Kit enables selective amplification of mitochondrial DNA from total DNA samples, without the need for prior mitochondrial DNA isolation. This kit provides DNA polymerase, buffers, and reagents for specific and uniform whole genome amplification from small samples of human and non-human mitochondrial DNA in total DNA samples. For amplification of non-human mtDNA, the REPLI-g Human mt Primer Mix needs to be substituted with an appropriate primer mix suitable for that species (not provided with the kit). The REPLI-g Mitochondrial DNA Kit is based on the multiple displacement amplification technology (MDA), that enriches for mitochondrial DNA with minimal contamination from nuclear DNA, thus avoiding the need for time-consuming isolation of mitochondrial DNA and increasing the sensitivity of downstream assays.