We applied DArTseq, and more recently DArTseqLD, to many hundreds of organisms for a wide range of applications, especially in genetic diversity characterisation and marker discovery area.
However, there are many applications which demand genotyping using selected marker set rather than randomly distributed markers reported by DArTseq/DArTseqLD methods. We have developed three technologies to offer the capacity to genotype materials with specific markers (mainly SNPs). These technologies vary in the marker density capacity as well as the initial cost of development, therefore we select the most appropriate technology for specific clients’ needs. Because the assays are always developed for specific material and marker sets we do not offer them to general public- you will not see them in our online ordering system unless you are representing the client for whom the assay was developed. Please contact us if you want to discuss your needs for specific marker characterisation.
Thanks to a number of technologies, we have developed we can offer a very cost effective solution to practically any application that requires anything from as few as a dozen of markers up to at least 10,000 markers.
DArTcap is used in similar applications as DArTseqLD, but it applies a selective step after complexity reduction to genotype specific markers from DArTseq representations. This selection is achieved with the use of the nucleic acid “capture probes” that bind to restriction fragments in the representations carrying the specific DArTseq markers. The restriction fragments are highly enriched in this capture step and most of the sequences obtained from the sequencing of the captured fragments represent the targeted markers. DArTcap is fairly flexible in terms of the number of markers that can be selected, but in most cases we target from a few hundreds to a few thousands markers. DArTcap is particularly useful in polyploids as the initial complexity reduction step (DArTseq library construction) prior to capture step would have already selected only one of the homeologous sequences at most loci. This feature enables highly effective sub-genome specific genotyping and obtaining co-dominant SNPs even in highly polyploid species with little or no genome sequence information.
DArTag method is targeting similar number of markers as DArTcap (many hundreds to thousands), but is not restricted to markers identified by DArTseq platform. Any SNP (or a small indel) can be targeted using DArTag if there is some genomic sequence available around the variant base/indel. In the first step of DArTag special molecular probes select the small target regions containing sequence variants. In the second step the targeted regions are amplified and, in parallel, the sample-specific barcode (a short stretch of sequence) is attached. The libraries generated in that manner are sequenced on the NGS equipment and resulting sequences processed using DArT PL’s proprietary pipeline.
DArTmp is used in applications requiring relatively small number of markers, but it can effectively process a few hundred SNPs in a single assay. There is no minimal number of markers that are required for the assay and DArT P/L has been delivering services with as few as 10 markers in one assay. However, the advantage of DArTmp compared to any single marker assay platforms is its ability to be increasingly fast as the loci it targets increases. The DArTmp assay is very similar in terms of requirements and the process to the DArTag assay as it also targets a short stretch of sequence around the sequence variant of interest. The main difference is the way the selection of the target sequence happens in the first step of the assay. In DArTmp a pair of oligonucleotide primers are used instead of a single probe applied in DArTag. All the consecutive steps in the lab and the whole analytical part are the same for DArTmp and DArTag. As DArTmp is targeting the smallest number of markers, the initial investment in assay development and oligo synthesis is small, enabling rapid adoption of this method to a number of applications requiring only modest number of markers.