MLST (Multilocus Sequence Typing) is a method of genetic typing of organisms based on determining the nucleotide sequence of a specific set of their genes (loci). This method was first proposed in 1998 for fast and reliable typing of pathogenic bacteria but can be successfully applied to any haploid organisms. In addition, to date, the method has been adapted for typing diploid organisms.
How does MLST work?
The method is based on the establishment of the nucleotide sequence of small fragments (about 500 pairs of nucleotides) of a number of genes and the subsequent comparison of the corresponding sequences in different organisms. In multilocus sequencing, scientists usually analyze the so-called housekeeping genes, which are necessary for the course of the reactions of the main metabolism, and therefore are present in all organisms. These genes, due to their exceptional importance for the viability of the organism, are characterized by a relatively low rate of accumulation of mutations, many of which are selectively neutral. In this regard, the comparison of the nucleotide sequences of such genes makes it relatively easy to establish the degree of the phylogenetic relationship between populations and systematize them. The number of loci analyzed in each particular study may be different, but most often it is 7-8. This amount provides sufficient resolution of the method and does not require too much labor, time and money for analysis.
Technically multilocus sequencing consists of several stages. After collecting the samples of microorganisms that need to be analyzed, DNA is isolated from them and the regions of certain genes are amplified by polymerase chain reaction using suitable primers. The nucleotide sequence of the amplified regions is then analyzed using automatic sequencers. The data obtained using special programs are compared with those available in the databases and conclusions are drawn about the status of specific genes in the studied populations. In the future, these data are used for epidemiological and population studies.
Advantages over other methods
Multilocus sequencing has several advantages over other older methods. One of them is a high resolution of the method: DNA sequencing makes it possible to identify more allelic variants than, for example, an electrophoretic study of the corresponding proteins, which was widely used previously. Secondly, the sequencing procedure is much easier to standardize than the conditions of many other typing methods, which facilitates the exchange of data between different laboratories, including via the Internet.