We present a survey of single cell whole genome amplification wga methods including degenerate oligonucleotide primed polymerase chain reaction dop pcr multiple displacement amplification mda and multiple annealing and looping based amplification cycles malbac.
Single cell genome amplification.
Multiple displacement amplification mda is a widely used technique enabling amplifying femtograms of dna from bacterium to micrograms for the use of sequencing.
Wga is achieved by pcr amplification of the library molecules using universal.
At very low levels of input dna successful amplification relies on reagent quality.
Depending on the method used the rate of artifact formation allelic dropout and sequence coverage over the genome may differ significantly.
Consequently accurate whole genome amplification wga of single cell dna is required for reliable genetic analysis e g ngs.
Single cell whole genome amplification and sequencing.
However current whole genome amplification wga methods are limited by low accuracy of copy number variation cnv detection and low amplification fidelity.
Single cell genomiphi dna amplification kit has been optimized to wholly amplify genomic dna from as little as a single cell generating micrograms of high quality dna for use in downstream applications.
Affiliation 1 biodynamic optical.
1 the kit includes a robust optimized cell lysis protocol that is incorporated into the.
Epub 2015 jun 11.
Here we report an improved single cell wga method linear amplification via trans.
Results the largest difference between the evaluated protocols was observed when analyzing the target coverage and read depth.
In contrast a deep analysis of the genome currently requires a few hundred nanograms up to micrograms of genomic dna for library formation necessary for ngs sequencing or labeling protocols e g microarrays.
Genomeplex single cell whole genome amplification kit utilizes a proprietary technology based on random fragmentation of genomic dna and conversion of the resulting small fragments to pcr amplifiable library molecules flanked by universal priming sites.
Single cell genomics is important for biology and medicine.
A list of more than 100 different single cell omics methods have been published.
Reagents required for mda reactions include.
Random primers and dna polymerase from bacteriophage phi29.
Whole genome amplification from a single cell is now possible with our optimized genomeplex single cell whole genome amplification kit the single cell procedure differs very little from the previously described genomeplex system but for three procedural changes.
1 pg of e coli dna has been amplified with either the repli g single cell kit or a kit based on malbac technology library prep and miseq sequencing similar data analysis on clc workbench.
The key parameters to characterize the performance of these methods are defined including genome coverage uniformity.
Methodology and applications annu rev genomics hum genet.
Single cell genomiphi has been developed with this in mind.
