Marker assisted selection or marker aided selection (MAS) is an indirect selection process where a
trait of interest is selected based on a
marker (
morphological,
biochemical or
DNA/
RNA variation) linked to a trait of interest (e.g. productivity, disease resistance, abiotic stress tolerance, and quality), rather than on the trait itself.
[Ribaut, J.-M. et al., Genetic basis of physiological traits. In Application of Physiology in Wheat Breeding, CIMMYT, Mexico, 2001.][Ribaut, J.-M. and Hoisington, D. A., Marker assisted selection: new tools and strategies. Trends Plant Sci., 1998, 3, 236–239.][Rosyara, U.R. 2006. REQUIREMENT OF ROBUST MOLECULAR MARKER TECHNOLOGY FOR PLANT BREEDING APPLICATIONS. Journal of Plant Breed. Gr. 1: 67 – 72]
click to download
/ref> This process has been extensively researched and proposed for plant
Plants are predominantly photosynthetic eukaryotes of the kingdom Plantae. Historically, the plant kingdom encompassed all living things that were not animals, and included algae and fungi; however, all current definitions of Plantae exclu ...
and animal breeding
Animal breeding is a branch of animal science that addresses the evaluation (using best linear unbiased prediction and other methods) of the genetic value (estimated breeding value, EBV) of livestock. Selecting for breeding animals with superior E ...
.
For example, using MAS to select individuals with disease resistance involves identifying a marker allele
An allele (, ; ; modern formation from Greek ἄλλος ''állos'', "other") is a variation of the same sequence of nucleotides at the same place on a long DNA molecule, as described in leading textbooks on genetics and evolution.
::"The chro ...
that is linked with disease resistance rather than the level of disease resistance. The assumption is that the marker associates at high frequency with the gene
In biology, the word gene (from , ; "... Wilhelm Johannsen coined the word gene to describe the Mendelian units of heredity..." meaning ''generation'' or ''birth'' or ''gender'') can have several different meanings. The Mendelian gene is a b ...
or quantitative trait locus
A quantitative trait locus (QTL) is a locus (section of DNA) that correlates with variation of a quantitative trait in the phenotype of a population of organisms. QTLs are mapped by identifying which molecular markers (such as SNPs or AFLPs) c ...
(QTL) of interest, due to genetic linkage (close proximity, on the chromosome, of the marker locus and the disease resistance-determining locus). MAS can be useful to select for traits that are difficult or expensive to measure, exhibit low heritability and/or are expressed late in development. At certain points in the breeding process the specimens are examined to ensure that they express the desired trait.
Marker types
The majority of MAS work in the present era uses DNA-based markers. However, the first markers that allowed indirect selection of a trait of interest were morphological markers. In 1923, Karl Sax first reported association of a simply inherited genetic marker A genetic marker is a gene or DNA sequence with a known location on a chromosome that can be used to identify individuals or species. It can be described as a variation (which may arise due to mutation or alteration in the genomic loci) that can be ...
with a quantitative trait in plants when he observed segregation of seed size associated with segregation for a seed coat color marker in beans ('' Phaseolus vulgaris'' L.). In 1935, J. Rasmusson demonstrated linkage of flowering time (a quantitative trait) in peas with a simply inherited gene for flower color.
Markers may be:
* Morphological – These were the first markers loci available that have an obvious impact on the morphology of plants. These markers are often detectable by eye, by simple visual inspection. Examples of this type of marker include the presence or absence of an awn, leaf sheath coloration, height, grain color, aroma of rice
Rice is the seed of the grass species '' Oryza sativa'' (Asian rice) or less commonly ''Oryza glaberrima'' (African rice). The name wild rice is usually used for species of the genera '' Zizania'' and '' Porteresia'', both wild and domesticat ...
etc. In well-characterized crops like maize
Maize ( ; ''Zea mays'' subsp. ''mays'', from es, maíz after tnq, mahiz), also known as corn (North American and Australian English), is a cereal grain first domesticated by indigenous peoples in southern Mexico about 10,000 years ago. The ...
, tomato
The tomato is the edible berry of the plant ''Solanum lycopersicum'', commonly known as the tomato plant. The species originated in western South America, Mexico, and Central America. The Mexican Nahuatl word gave rise to the Spanish word ...
, pea, barley
Barley (''Hordeum vulgare''), a member of the grass family, is a major cereal grain grown in temperate climates globally. It was one of the first cultivated grains, particularly in Eurasia as early as 10,000 years ago. Globally 70% of barley p ...
or wheat
Wheat is a grass widely cultivated for its seed, a cereal grain that is a worldwide staple food. The many species of wheat together make up the genus ''Triticum'' ; the most widely grown is common wheat (''T. aestivum''). The archaeologi ...
, tens or hundreds of genes that determine morphological traits have been mapped to specific chromosome locations.
*Biochemical – A protein that can be extracted and observed; for example, isozyme In biochemistry, isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. Isozymes usually have different kinetic parameters (e.g. dif ...
s and storage protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, res ...
s.
* Cytological – Cytological markers are chromosomal
A chromosome is a long DNA molecule with part or all of the genetic material of an organism. In most chromosomes the very long thin DNA fibers are coated with packaging proteins; in eukaryotic cells the most important of these proteins are ...
features that can be identified through microscopy. These generally take the form of chromosome bands, regions of chromatin
Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in r ...
that become impregnated with specific dyes used in cytology
Cell biology (also cellular biology or cytology) is a branch of biology that studies the structure, function, and behavior of cells. All living organisms are made of cells. A cell is the basic unit of life that is responsible for the living an ...
. The presence or absence of a chromosome band can be correlated with a particular trait, indicating that the locus responsible for the trait is located within or near (tightly linked) to the banded region. Morphological and cytological markers formed the backbone of early genetic studies in crops such as wheat and maize.
*DNA-based- Including microsatellites
A microsatellite is a tract of repetitive DNA in which certain DNA motifs (ranging in length from one to six or more base pairs) are repeated, typically 5–50 times. Microsatellites occur at thousands of locations within an organism's genome. ...
(also known as short tandem repeats, STRs, or simple sequence repeats, SSRs), restriction fragment length polymorphism (RFLP), random amplification of polymorphic DNA
Random amplification of polymorphic DNA (RAPD), pronounced "rapid", is a type of polymerase chain reaction (PCR), but the segments of DNA that are amplified are random. The scientist performing RAPD creates several arbitrary, short primers (10- 1 ...
(RAPD), amplified fragment length polymorphism
AFLP-PCR or just AFLP is a PCR-based tool used in genetics research, DNA fingerprinting, and in the practice of genetic engineering. Developed in the early 1990s by KeyGene, AFLP uses restriction enzymes to digest genomic DNA, followed by liga ...
(AFLP), and single nucleotide polymorphisms (SNPs).
Positive and negative selectable markers
The following terms are generally less relevant to discussions of MAS in plant and animal breeding, but are highly relevant in molecular biology research:
*Positive selectable markers are selectable markers that confer selective advantage to the host organism. An example would be antibiotic resistance, which allows the host organism to survive antibiotic selection.
*Negative selectable markers are selectable markers that eliminate or inhibit growth of the host organism upon selection. An example would be thymidine kinase
Thymidine kinase is an enzyme, a phosphotransferase (a kinase): 2'-deoxythymidine kinase, ATP-thymidine 5'-phosphotransferase, EC 2.7.1.21. It can be found in most living cells. It is present in two forms in mammalian cells, TK1 and TK2. Cert ...
, which makes the host sensitive to ganciclovir
Ganciclovir, sold under the brand name Cytovene among others, is an antiviral medication used to treat cytomegalovirus (CMV) infections.
Ganciclovir was patented in 1980 and approved for medical use in 1988.
Medical use
Ganciclovir is indicated ...
selection.
A distinction can be made between selectable markers (which eliminate certain genotypes from the population) and screenable markers (which cause certain genotypes to be readily identifiable, at which point the experimenter must "score" or evaluate the population and act to retain the preferred genotypes). Most MAS uses screenable markers rather than selectable markers.
Gene vs marker
The gene of interest directly causes production of protein(s) or RNA that produce a desired trait or phenotype, whereas markers (a DNA sequence or the morphological or biochemical markers produced due to that DNA) are genetically linked to the gene of interest. The gene of interest and the marker tend to move together during segregation of gametes due to their proximity on the same chromosome and concomitant reduction in recombination (chromosome crossover events) between the marker and gene of interest. For some traits, the gene of interest has been discovered and the presence of desirable alleles can be directly assayed with a high level of confidence. However, if the gene of interest is not known, markers linked to the gene of interest can still be used to select for individuals with desirable alleles of the gene of interest. When markers are used there may be some inaccurate results due to inaccurate tests for the marker. There also can be false positive results when markers are used, due to recombination between the marker of interest and gene (or QTL). A perfect marker would elicit no false positive results. The term 'perfect marker' is sometimes used when tests are performed to detect a SNP or other DNA polymorphism in the gene of interest, if that SNP or other polymorphism is the direct cause of the trait of interest. The term 'marker' is still appropriate to use when directly assaying the gene of interest, because the test of genotype is an indirect test of the trait or phenotype of interest.
Important properties of ideal markers for MAS
An ideal marker:
*Has easy recognition of phenotypes - ideally all possible phenotypes (homo
''Homo'' () is the genus that emerged in the (otherwise extinct) genus '' Australopithecus'' that encompasses the extant species ''Homo sapiens'' ( modern humans), plus several extinct species classified as either ancestral to or closely relat ...
- and heterozygotes
Zygosity (the noun, zygote, is from the Greek "yoked," from "yoke") () is the degree to which both copies of a chromosome or gene have the same genetic sequence. In other words, it is the degree of similarity of the alleles in an organism.
Mo ...
) from all possible alleles
*Demonstrates measurable differences in expression between trait types or gene of interest alleles, early in the development of the organism
*Testing for the marker does not have variable success depending on the allele at the marker locus or the allele at the target locus (the gene of interest that determines the trait of interest).
*Low or null interaction among the markers allowing the use of many at the same time in a segregating population
*Abundant in number
* Polymorphic
Drawbacks of morphological markers
Morphological markers are associated with several general deficits that reduce their usefulness including:
*the delay of marker expression until late into the development of the organism
*allowing dominance to mask the underlying genetics
* pleiotropy, which does not allow easy and parsimonious inferences to be drawn from one gene to one trait
*confounding effects of genes unrelated to the gene or trait of interest but which also affect the morphological marker ( epistasis)
*frequent confounding effects of environmental factors which affect the morphological characteristics of the organism
To avoid problems specific to morphological markers, DNA-based markers have been developed. They are highly polymorphic, exhibit simple inheritance (often codominant), are abundant throughout the genome, are easy and fast to detect, exhibit minimum pleiotropic effects, and detection is not dependent on the developmental stage of the organism. Numerous markers have been mapped to different chromosomes in several crops including rice, wheat, maize, soybean and several others, and in livestock such as cattle, pigs and chickens. Those markers have been used in diversity analysis, parentage detection, DNA fingerprinting, and prediction of hybrid performance. Molecular markers are useful in indirect selection processes, enabling manual selection of individuals for further propagation.
Selection for major genes linked to markers
'Major genes' that are responsible for economically important characteristics are frequent in the plant kingdom. Such characteristics include disease resistance, male sterility, self-incompatibility, and others related to shape, color, and architecture of whole plants and are often of mono- or oligogenic in nature. The marker loci that are tightly linked to major genes can be used for selection and are sometimes more efficient than direct selection for the target gene. Such advantages in efficiency may be due for example, to higher expression of the marker mRNA in such cases that the marker is itself a gene. Alternatively, in such cases that the target gene of interest differs between two alleles by a difficult-to-detect single nucleotide polymorphism, an external marker (be it another gene or a polymorphism that is easier to detect, such as a short tandem repeat
A microsatellite is a tract of repetitive DNA in which certain DNA motifs (ranging in length from one to six or more base pairs) are repeated, typically 5–50 times. Microsatellites occur at thousands of locations within an organism's genome. ...
) may present as the most realistic option.
Situations that are favorable for molecular marker selection
There are several indications for the use of molecular markers in the selection of a genetic trait.
Situations such as:
*The selected character is expressed late in plant development, like fruit and flower features or adult characters with a juvenile period (so that it is not necessary to wait for the organism to become fully developed before arrangements can be made for propagation)
*The expression of the target gene is recessive (so that individuals which are heterozygous positive for the recessive allele can be crossed to produce some homozygous offspring with the desired trait)
*There are special conditions for expression of the target gene(s), as in the case of breeding for disease and pest resistance (where inoculation with the disease or subjection to pests would otherwise be required). Sometimes inoculation methods are unreliable and sometimes field inoculation with the pathogen is not even allowed for safety reasons. Moreover, sometimes expression is dependent on environmental conditions.
*The phenotype is affected by two or more unlinked genes (epistatis). For example, selection for multiple genes which provide resistance against diseases or insect pests for gene pyramiding.
The cost of genotyping
Genotyping is the process of determining differences in the genetic make-up (genotype) of an individual by examining the individual's DNA sequence using biological assays and comparing it to another individual's sequence or a reference sequence. ...
(for example, the molecular marker assays needed here) is decreasing thus increasing the attractiveness of MAS as the development of the technology continues. (Additionally, the cost of phenotyping performed by a human is a labor burden
Labor burden is the actual cost of a company to have an employee, aside from the salary the employee earns. Labor burden costs include benefits that a company must, or chooses to, pay for employees included on their payroll. These costs include bu ...
, which is higher in a developed country and increasing in a developing country.)
Steps for MAS
Generally the first step is to map
A map is a symbolic depiction emphasizing relationships between elements of some space, such as objects, regions, or themes.
Many maps are static, fixed to paper or some other durable medium, while others are dynamic or interactive. Although ...
the gene or quantitative trait locus
A quantitative trait locus (QTL) is a locus (section of DNA) that correlates with variation of a quantitative trait in the phenotype of a population of organisms. QTLs are mapped by identifying which molecular markers (such as SNPs or AFLPs) c ...
(QTL) of interest first by using different techniques and then using this information for marker assisted selection. Generally, the markers to be used should be close to gene of interest (<5 recombination unit or cM) in order to ensure that only minor fraction of the selected individuals will be recombinants. Generally, not only a single marker but rather two markers are used in order to reduce the chances of an error due to homologous recombination. For example, if two flanking markers are used at same time with an interval between them of approximately 20cM, there is higher probability (99%) for recovery of the target gene.
QTL mapping techniques
In plants QTL mapping is generally achieved using bi-parental cross populations; a cross between two parents which have a contrasting phenotype for the trait of interest are developed. Commonly used populations are near isogenic lines (NILs), recombinant inbred lines (RILs), doubled haploids (DH), back cross and F2. Linkage between the phenotype and markers which have already been mapped is tested in these populations in order to determine the position of the QTL. Such techniques are based on linkage and are therefore referred to as "linkage mapping
Genetic linkage is the tendency of DNA sequences that are close together on a chromosome to be inherited together during the meiosis phase of sexual reproduction. Two genetic markers that are physically near to each other are unlikely to be separ ...
".A
Single step MAS and QTL mapping
In contrast to two-step QTL mapping and MAS, a single-step method for breeding typical plant populations has been developed.[Rosyara, U. R.; K.L. Maxson-Stein; K.D. Glover; J.M. Stein; J.L. Gonzalez-Hernandez. 2007. Family-based mapping of FHB resistance QTLs in hexaploid wheat. Proceedings of National Fusarium head blight forum, 2007, Dec 2–4, Kansas City, MO.][Rosyara U.R., J.L. Gonzalez-Hernandez, K.D. Glover, K.R. Gedye and J.M. Stein. 2009. Family-based mapping of quantitative trait loci in plant breeding populations with resistance to Fusarium head blight in wheat as an illustratio]
Theoretical Applied Genetics 118:1617–1631
/ref>
In such an approach, in the first few breeding cycles, markers linked to the trait of interest are identified by QTL mapping and later the same information is used in the same population. In this approach, pedigree structure is created from families that are created by crossing number of parents (in three-way or four way crosses). Both phenotyping and genotyping is done using molecular markers mapped the possible location of QTL of interest. This will identify markers and their favorable alleles. Once these favorable marker alleles are identified, the frequency of such alleles will be increased and response to marker assisted selection is estimated. Marker allele(s) with desirable effect will be further used in next selection cycle or other experiments.
High-throughput genotyping techniques
Recently high-throughput genotyping techniques are developed which allows marker aided screening of many genotypes. This will help breeders in shifting traditional breeding to marker aided selection. One example of such automation is using DNA isolation robots, capillary electrophoresis and pipetting robots.
One recent example of capllilary system is Applied Biosystems 3130 Genetic Analyzer. This is the latest generation of 4-capillary electrophoresis instruments for the low to medium throughput laboratories.
High-throughput MAS is needed for crop breeding because current techniques are not cost effective. Arrays have been developed for rice by Masouleh et al 2009; wheat by Berard et al 2009, Bernardo et al 2015, and Rasheed et al 2016; legumes by Varshney et al 2016; and various other crops, but all of these have also problems with customization, cost, flexibility, and equipment costs.
Use of MAS for backcross breeding
A minimum of five or six-backcross
Backcrossing is a crossing of a hybrid with one of its parents or an individual genetically similar to its parent, to achieve offspring with a genetic identity closer to that of the parent. It is used in horticulture, animal breeding, and product ...
generations are required to transfer a gene of interest from a donor (may not be adapted) to a recipient (recurrent – adapted cultivar). The recovery of the recurrent genotype can be accelerated with the use of molecular markers. If the F1 is heterozygous for the marker locus
Locus (plural loci) is Latin for "place". It may refer to:
Entertainment
* Locus (comics), a Marvel Comics mutant villainess, a member of the Mutant Liberation Front
* ''Locus'' (magazine), science fiction and fantasy magazine
** ''Locus Award' ...
, individuals with the recurrent parent allele(s) at the marker locus in first or subsequent backcross generations will also carry a chromosome tagged by the marker.
Marker assisted gene pyramiding
Gene pyramiding has been proposed and applied to enhance resistance to disease and insects by selecting for two or more than two genes at a time. For example, in rice such pyramids have been developed against bacterial blight and blast. The advantage of use of markers in this case allows to select for QTL-allele-linked markers that have same phenotypic effect.
MAS has also been proved useful for livestock
Livestock are the domesticated animals raised in an agricultural setting to provide labor and produce diversified products for consumption such as meat, eggs, milk, fur, leather, and wool. The term is sometimes used to refer solely to animal ...
improvement.
A coordinated effort to implement wheat (''Triticum turgidum
Durum wheat (), also called pasta wheat or macaroni wheat (''Triticum durum'' or ''Triticum turgidum'' subsp. ''durum''), is a tetraploid species of wheat. It is the second most cultivated species of wheat after common wheat, although it represen ...
'' and ''Triticum aestivum
Common wheat (''Triticum aestivum''), also known as bread wheat, is a cultivated wheat species. About 95% of wheat produced worldwide is common wheat; it is the most widely grown of all crops and the cereal with the highest monetary yield.
Ta ...
)'' marker assisted selection in the U.S. as well as a resource for marker assisted selection exists at the Wheat CAP (Coordinated Agricultural Project) website.
See also
* Association mapping In genetics, association mapping, also known as " linkage disequilibrium mapping", is a method of mapping quantitative trait loci (QTLs) that takes advantage of historic linkage disequilibrium to link phenotypes (observable characteristics) to geno ...
* Family based QTL mapping
Quantitative trait loci mapping or QTL mapping is the process of identifying genomic regions that potentially contain genes responsible for important economic, health or environmental characters. Mapping QTLs is an important activity that plant ...
* Genomics of domestication
* History of plant breeding
* Molecular breeding
Molecular breeding is the application of molecular biology tools, often in plant breedingVoosen P (2009Molecular Breeding Makes Crops Hardier and More Nutritious Markers, knockouts and other technical advances improve breeding without modifying gen ...
* Nested association mapping Nested association mapping (NAM) is a technique designed by the labs of Edward BucklerJames Holland anfor identifying and dissecting the genetic architecture of complex traits in corn (''Zea mays''). It is important to note that nested association ...
* QTL mapping
A quantitative trait locus (QTL) is a locus (section of DNA) that correlates with variation of a quantitative trait in the phenotype of a population of organisms. QTLs are mapped by identifying which molecular markers (such as SNPs or AFLPs) co ...
*
* Smart breeding
References
Further reading
review application of MAS in crop improvement
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Plant Breeding and Genomics
{{DEFAULTSORT:Marker Assisted Selection
Genetics
de:Marker Assisted Selection