Also known as marker molecules or genetic markers, they are used to mark the position of a specific gene under investigation, or draw attention to the inheritance of a characteristic.
They have proven to be an essential tool for geneticists and have found essential applications in genetic engineering, paternity testing and the identification of deadly diseases.
The molecular markers are DNA fragments that are associated with a particular region of the genome. They can take the form of short DNA sequences, such as a sequence surrounding a single nucleotide polymorphism, where a single base pair change occurs.
They can also take the form of longer DNA sequences, such as microsatellites, which are 10 to 60 base pairs long.
Classes of molecular markers
Restriction fragment length polymorphisms are molecular markers that are used to track a particular DNA sequence as it passes between cells.
It is one of the most common types of molecular markers and is based on the hybridization of cloned DNA to DNA fragments. They are specific for a single combination of clone or restriction enzyme.
Molecular markers of randomly amplified polymorphic DNA are generally used in plant breeding and are based on the cloning of the polymer chain reaction gene from random locations of a plant genome.
Molecular markers of isoenzymes are used to label proteins. They are designed to identify enzymes that differ in amino acid sequences but that catalyze the same amino acid reaction.
Some common types of genetic markers are:
- RFLP (or restriction fragment length polymorphism).
- AFLP (or fragment length polymorphism amplified.
- RAPD (or random amplification of polymorphic DNA).
- VNTR (or variable number repetition).
- Microsatellite polymorphism.
- SNP (or single nucleotide polymorphism).
- STR (or short repeat).
- SFP (or single-feature polymorphism) DArT (or diversity matrix technology).
They can be categorized as dominant or co-dominant. The dominant markers allow to analyze many loci at the same time, for example, RAPD.
A primer that amplifies a dominant marker could be amplified at many loci in a DNA sample with a PCR reaction. Co-dominant markers analyze one locus at a time.
An amplifying primer of a co-dominant marker would produce a specific product.
Uses of molecular markers
Geneticists use molecular markers to examine the relationship between hereditary diseases and their causes.
They can indicate the location of a specific mutation of a gene that can result in a damaged protein and have been used to identify diseases such as sickle cell anemia and Huntington’s disease.
Molecular markers can also have agricultural applications, such as marker-assisted breeding, in paternity tests and in the identification of plant varieties by identifying the identity, purity and stability of a plant.
Molecular markers are used in genetic engineering to mark a place where defective and mutated proteins have been replaced by others that function correctly.
This is done by replacing the damaged DNA sequence with an identical but well-functioning sequence transplanted from another location.
Characteristics of an ideal molecular marker
An ideal molecular marker must have all or at least some of the following characteristics:
- It should be easily available.
- It must be easy to analyze.
- It should not be time consuming.
- It must be highly reproducible.
- It must be phenotypically neutral.
- It can produce interchangeable data.
- It should show polymorphism.
- It must show the inheritance of codomination to allow discrimination between homozygotes and heterozygotes.
- It must be non-epistatic.
A series of different molecular markers of the system, which was available during the last two decades, can be classified in general into three classes:
- The first generation molecular markers, including the RFLPs, the RAPDs and their modifications.
- The second generation molecular marker that includes SSR, AFLP and its modified forms.
- Molecular markers of third generation: environmentally sound technologies, SNP and many more.
Several approaches are available to identify markers related to the trait of interest. Molecular marker maps are now available for the most important species.
Molecular markers have been considered as a tool for a large number of applications ranging from the localization of a gene to the improvement of species through selection assisted by markers.
They have also become extremely popular for phylogenetic analysis that adds new dimensions to evolutionary theories in plant and animal research.
Molecular markers and their reproducibility
Reproducibility is always an important property of markers, but even more important with collaborative projects, which involves the generation of data by different laboratories whose results must be assembled.
To obtain reproducible results, the extraction of high quality purified DNA is a prerequisite for most marker techniques. For example, degraded and / or unpurified DNA can affect the amplification or restriction of DNA, resulting in nonspecific polymorphisms.
Even when purified and high molecular weight DNA is used, RAPD often do not show reproducible results.
This is because the RAPD primers are very short (10 bp), which can result in alterations in their annealing behavior to the template DNA and the resulting band profiles as a result of small deviations under experimental conditions.
Therefore, highly standardized experimental procedures are required when using RAPD markers. This implies the need to include repeated samples and also the inclusion of reference genotypes that represent bands of known size.
Problems with reproducibility in the RAPD analysis could be overcome by focusing on the mapped markers for which their inheritance has already been verified.