What is Diversity Array Technology (DArT)?

Introduction

Genetic diversity can be defined as the variation in the traits that are passed from one generation to another within a species. More is the genetic diversity more is the variation in the traits. This genetic variation can be utilized to determine the sequence information of an individual. Utilization of genetic diversity to analyse and create genetic map is called diversity arrays technology (DArT).

DArT is a comparatively newer approach to assess polymorphism which has been developed due to limitations of existing molecular marker techniques. This technique has been successfully utilized in the plant breeding programmes.

What is Diversity array technology (DArT)

DArT is a novel technology which does not dependent on the sequence of organism, high-throughput technique and is capable of detecting many markers in one experiment. It provides a cheaper, more practical and fingerprinting of whole genome approach to stud and map the diversity among the individuals. This technique was developed by Kaiman Peng, Andrzej Killian, David Feinstein and Damian Jaccoud in 2001.

This technique a little amount of initial DNA can be used to analyse large amount of samples. Also, it is faster and far more cost effective as compared to other molecular markers like single nucleotide polymorphism (SNPs).

Steps Involved in DArT

DArT mainly involves three steps, they are −

• Reduction of complexity.

• Representation of genomic DNA.

• Assay.

Reduction of Complexity

In this step certain restriction enzymes are used to cleave large genomic DNA from a specific species of interest into numerous smaller fragments as genomic DNA is relatively complex and cannot be managed easily.

• As the sample to be analysed contain elevated polymorphism therefore the coupling between restriction enzyme to be selected and primer for annealing becomes crucial.

• The restriction enzyme used should be highly specific for the non-repetitive sequences, nonmethylated genome of the species. One of the most commonly used restriction enzymes with these qualities is PstI which is a type II restriction endonuclease and has been widely used in cloning, RFLP and genotyping.

Representation of Genomic DNA

After the digestion of genomic DNA using specific restriction enzymes, this step involves the selection of the digest that contains the largest quantity of polymorphism in the gene pool, which is of significance.

• Since these smaller fragments act as the representatives of the much larger genomic DNA, they are known as representations.

• It is also advisable to not select the repetitive sequences as they show lowest amount of polymorphism or variations. Accurate results are obtained when non-repetitive sequences are in abundance.

DArT

After representations are produced double stranded DNA is produced by the joining of the digested fragments using T4 ligase a small amount of which is diluted and amplified using polymerase chain reaction.

• It is important to use restriction site complementary primers and he variant of Taq polymerase which cannot be inhibited during polymerase chain reaction.

• The product is then mixed with the representation and then it is inserted to a vector. This vector is then transformed into a suitable host by either chemical method or by electrical shock method.

• The transformed cells are then incubated in a suitable medium and allowed to grow and selection is done based on antibiotic resistance in X-gal medium.

• Then inserts are amplified by polymerase chain reaction and then product is inserted in the microarray slides. The slides are centrifuged to isolate the insert which is then purified.

• One more technique which uses software and fluorescent probes for the detection of polymorphism and that software is known as DArTsoft.

Advantages of DArT

• The most important advantage of this technique is that it does not dependent on the sequence of the DNA which make it available for many species.

• It is the user of this technique who decides the scope of the genetic analysis and it can be used for certain species and can be expanded to the wild varieties also.

• This technology has a very high throughput it means that it has the ability to identify numerous polymorphic markers in parallel. The cost of this technique is very less as compared to others and this can be reduced further.

Applications of DArT

• Like RFLP, SNPs, AFLP etc. this can also be used as genetic marker. With this technique genomic profiles of any crop can be obtained at low cost.

• They can also be used for the collection of germ plasm which can be used for recognition and management of biodiversity.

• This method is so fast that it allows plant breeder to map quantitative trait loci (QTL) in less time which allow them to emphasize on more important factors in plant breeding. Like reliable and precise phenotyping.

• Since it has wide application in agricultural biotechnology, so it an be used in developing countries to increase the pace of plant breeding and maintaining the biodiversity.

Conclusion

DArT is relatively unique tool which helps in DNA profiling without the requirement of genome sequence. It is a proven tool in the field of plant breeding with much lesser cost. DArT has been successfully implemented in some cereals like wheat, barley and rice, also more research work is going on to develop it in certain varieties of sorghum, chickpea and pigeon pea, which can have a greater impact on developing countries.