Developing Protocols for Animal Genebanks : Exploiting Limited Amounts of DNA By Whole Genome Amplification

BRC missions are to provide a secure storage and easy access to samples for users. Limiting stocks of DNA do not always allow supplying enough material to perform genomic analysis. The CRB-Anim infrastructure project has supported technological developments for the exploitation of samples with limited DNA amounts.

The whole genome amplification (WGA) methods, based on the multiple displacement amplification approach (MDA) using Phi29 polymerase, are effective strategies to generate high-quality genomic DNA (gDNA) in sufficient amounts for successful SNP high throughput genotyping. 3 MDA kits, namely Repli-g (QIAGEN), TruePrime (SYGNIS) and GenomiPhi (GE Healthcare) were tested with various starting quantities of gDNA or directly on small amounts of blood. To assess the amplification quality and to estimate error rates, genotyping was performed on Illumina Bovine EuroG10K v4 chips. The first part of the test consisted in gDNA amplification and the second in direct amplification after blood cell lysis. First, each genomic DNA sample (3 bulls, 10 ng and 1 ng) was amplified 3 times; second, each blood sample (2 bulls, 0.5 µl and 1 µl) was amplified in duplicate, using the 3 WGA kits. Genotypes were compared to those previously obtained with native unamplified DNA. Whatever input source, the call rate was equal to the one observed with native DNA and the error rate ranged from 0.01 % to 30% according to the quality of DNA input. The direct use of small amounts of blood provided more homogenous results than the amplification from residual DNA.