REPLI-g UltraFast Mini Kit

For ultrafast highly uniform whole genome amplification from small or precious samples

Fast 1–1.5 hour whole genome amplification
Obtain unlimited DNA for downstream applications
Reproducible amplification from a variety of starting materials
Highly stable DNA suitable for long-term storage

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.

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Product		Cat. no.	List price:
REPLI-g UltraFast Mini Kit (25) DNA Polymerase, Buffers, and Reagents for 25 x 20 μl ultrafast whole genome amplification reactions		150033	Inquire
REPLI-g UltraFast Mini Kit (100) DNA Polymerase, Buffers, and Reagents for 100 x 20 μl ultrafast whole genome amplification reactions		150035	Inquire

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The REPLI-g UltraFast Mini Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

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Purified genomic DNA procedure.|Consistent and high yields of amplified DNA.|

Amplification of genomic DNA using the REPLI-g UltraFast Mini Kit involves 3 basic steps. First, the sample undergoes gentle alkaline denaturation. Next, the sample is neutralized, and finally incubated with REPLI-g UltraFast DNA Polymerase at 30°C.|Genomic DNA samples (10 ng each) and non-template control (NTC) samples (containing no template DNA) were amplified using the REPLI-g UltraFast Mini Kit and the DNA yield was determined after various times. An average yield of 7 µg was obtained after amplification for 60 minutes, while 10–13 µg DNA was obtained after 90 minutes. Up to 5 µg DNA was obtained after just 45 minutes, allowing even faster downstream genetic analysis. No DNA was detected in the non-template control reactions.|

Performance

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.

Principle

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.

Procedure

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.

Applications

REPLI-g amplified genomic DNA can be used in a variety of downstream applications, including:

Genotyping (e.g., SNP, STR, microarray)
End-point PCR, quantitative real-time PCR
Sequencing
Comparative genome hybridization

Feature	Specifications
Amplification	Whole genomic DNA
Applications	Genotyping, microarray, PCR, real-time PCR
Denaturation step	Alkaline
Maximum input volume	10 ng DNA, 0.5 µl whole blood, ~300 cells/µl
Minimal pipetting volume needed	1 µl
Quality assessment	No
Reaction time	60–90 minutes
Reaction volume	20 µl
Samples per run (throughput)	Mid
Starting amount of DNA	>10 ng purified genomic DNA
Starting material	Genomic DNA, blood, cells
Technology	Multiple Displacement Amplification (MDA)
Yield	~10 µg

Brochures & Guides

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	Analyzing Genetic Differences - (EN) Second edition — innovative tools	Show details

FAQs

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	Can I reduce the amount of starting material below 10 ng when using the REPLI-g UltraFast Mini Kit? FAQ ID -1325	View
	Can I reduce the reaction time to e.g., 30 minutes, when using the REPLI-g UltraFast Mini Kit? FAQ ID -1326	View
	Why is there DNA in the no-template control reaction when using the standard REPLI-g procedure, but not when using the UltraFast procedure? FAQ ID -1327	View
	Will the REPLI-g UltraFast Mini Kit reaction always yield 10 µg DNA? FAQ ID -1328	View
	Do you have a protocol for cleanup of REPLI-g amplified DNA? FAQ ID -1545	View
	Can cDNA prepared with the QuantiTect Whole Transcriptome Kit be reampliied using REPLI-g Kits? FAQ ID -1611	View
	Where can I find background information and literature on Whole Genome Amplification with REPLI-g Kits? FAQ ID -1690	View
	What are possible reasons for reduced DNA yields with REPLI-g Kits? FAQ ID -2148	View
	Do you have data on the DNA yield at different incubation time for the Repli-G Kits? FAQ ID -3113	View
	What is REPLI-g whole genome amplification? FAQ ID -654	View
	Will REPLI-g work at high temperatures? FAQ ID -656	View
	What are the differences between MDA and DOP/PEP methods of Whole Genome Amplification? FAQ ID -665	View
	Does REPLI-g technology for whole genome amplification work with paraffin embedded samples? FAQ ID -673	View
	Why do I get amplification in a negative control DNA tube using the REPLI-g Kit for WGA? FAQ ID -675	View
	Is 2 kb the minimal gDNA fragment size for REPLI-g Whole Genome Amplification? FAQ ID -682	View
	What is the length of the MDA whole genome amplification product? FAQ ID -690	View
	What is the stability of the REPLI-g MDA product? FAQ ID -693	View
	How can I quantify the amount of REPLI-g DNA I have amplified? FAQ ID -694	View
	How can I determine the quality of my REPLI-g amplified products? FAQ ID -695	View
	Has anyone verified whole genome amplification accuracy with Sequencing? FAQ ID -701	View
	Are Centromeres and Telomeres amplified using REPLI-g WGA? FAQ ID -702	View
	What is the enzyme used in the REPLI-g reaction? FAQ ID -704	View
	Any data on the fidelity of the REPLI-g MDA technique? FAQ ID -707	View
	Can I purchase Phi29 DNA polymerase only? FAQ ID -708	View
	What are exo-resistant random hexamers used in the REPLI-g reaction? FAQ ID -710	View
	Can I use my own primers for REPLI-g WGA to amplify a specific chromosomal region? FAQ ID -712	View
	Will the random hexamers in the REPLI-g reaction interfere with downstream analysis? FAQ ID -713	View
	Can I use REPLI-g for SNP Genotyping? FAQ ID -700	View

Scientific Posters

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	A novel approach to whole genome amplification and labeling of DNA samples for copy number variation detection on BAC microarrays	Show details
	Comparison of genotyping consistency between genomic and whole-genome amplified DNA using the Illumina GoldenGate and Infinium-II assays	Show details
	SNP genotyping of saliva DNA using Affymetrix GeneChip targeted genotyping system	Show details
	(EN) - The impact of whole genome amplification on forensic testing	Show details

Application Notes

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	Genome-pULSe: Whole genome amplification & labeling for arrayCGH analysis - (EN)	Show details
	Oragene and REPLI-g Whole Genome Amplifi cation - (EN)	Show details

Kit Handbooks

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	(EN) - REPLI-g UltraFast Mini Handbook For fast whole genome amplification from purified genomic DNA, blood, and cells	Show details

Supplementary Protocols

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	Purification of REPLI-g amplified DNA using the QIAamp® DNA Mini Kit - (EN)	Show details
	Purification of DNA amplified using REPLI-g® Kits - (EN)	Show details

Safety Data Sheets

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	MSDS REPLI-g UltraFast Mini Kit (25)
	MSDS REPLI-g UltraFast Mini Kit (100)

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