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Hello. Molecular genetics 
influences new breeding methods.

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This lecture focuses on genetic 
markers and their applications.

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The lecture is part of Module 3, Animal Breeding. 
The creation of this presentation was supported by

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ERASMUS+ KA2 grant within the ISAGREED project, 
Innovation of content and structure of study

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programs in the field of management of animal 
genetic and food resources using digitization.

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With the development of molecular genetics since 
the 1970s, and especially molecular genetic

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methods working with DNA molecules, it is possible 
to identify real genetic variability (genotypes)

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using molecular genetic markers. This is used, 
for example, for mapping genes or regions in

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the genome (QTL) where genes for complex utility 
traits might be located. Thanks to these analyses,

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the use of this information in breeding 
is possible, specifically for improving

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estimates of genetic parameters (such as 
heritability) and breeding values, which leads

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to the efficiency of breeding through increased 
genetic gain and shortened generation interval.

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At the beginning (i.e. since the 1980s), 
the idea of using genetic markers as a

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selection criterion emerged, i.e. after 
identifying the genotype of individuals,

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subsequent selection of a genotype-appropriate 
parent. This is called marker-assisted selection

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(MAS). However, this approach is limited in 
its actual use. Such selection has been used,

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for example, for the stress gene in pigs (CRC 
gene) or BLAD in cattle. These were genes with

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monogenic inheritance, mutations of which 
caused a reduction in individual fitness.

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For utility, complex, quantitative traits, direct 
selection based on the genotype of a single gene

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is inappropriate, or multiple markers had to 
be included in statistical models and breeding

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values were estimated. If the variability in the 
causal gene was directly determined by a marker,

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it is called gene-assisted selection (GAS).
A great revolution in these approaches was the

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advent of new technologies for massive genome 
sequencing and identification of millions of

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markers (SNPs) and more precise determination 
of QTL regions and genes in the genome for

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quantitative traits. Since 2009, after sequencing 
the genomes of cattle, and gradually other

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economically important animal species, genomic 
selection has been gradually introduced. This is

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a method that incorporates genomic SNP markers 
(tens of thousands to hundreds of thousands)

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into a genomic relationship matrix and into 
equations such as BLUP (various variants) in

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order to estimate breeding values - 
GEBV (genomic estimated breeding value).

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This is again just one number that is 
easily usable in breeding practice.

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Because we use real genetic variability, breeding 
is significantly streamlined - costs are reduced,

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breeding value estimates are improved, 
and the generation interval is shortened.

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So what is a genetic molecular marker? It is a 
detectable polymorphism (multiple alleles) with a

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known position in the genome. There are three 
types of these markers: Type I are coding genes,

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so-called candidate genes (e.g. CRC gene, ESR 
in pigs). Type II markers are microsatellites,

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i.e. short tandemly repeating base sequences 
(STR) - found outside coding sequences.

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Type III markers are biallelic single nucleotide 
polymorphisms (SNPs) in coding or more commonly

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in non-coding intronic or intergenic regions. An 
example of one SNP is shown in the bottom image.

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The diagram shows polymorphism in 
SNPs for a single nucleotide pair,

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and polymorphism in microsatellites, 
which is caused by length variation of

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tandem repeats (especially dinucleotide 
repeats). SNPs usually have two alleles,

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while microsatellites can have up 
to twenty alleles in a population,

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although there are significantly fewer 
of them in genomes compared to SNPs.

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The main significance of a genetic marker is that 
it can be associated with phenotypic variability

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of an important production trait in breeding, even 
though it may not have a direct biological effect

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on the trait. Such a marker is then referred to 
as indirect. It is called indirect because it

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is in linkage (i.e. close proximity on the 
chromosome) with another gene (QTL region)

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that directly influences the trait. The marker's 
linkage disequilibrium with QTL is then utilized.

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The stronger the linkage, the more 
informative the marker is for breeding.

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An example of a SNP that is also a candidate 
gene is a SNP on the 4th chromosome,

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located in the leptin gene, where there is a 
polymorphism of adenine and guanine substitution.

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Furthermore, we can see that the mutation is 
in the coding sequence and causes a change in

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reading, substituting the 40th amino acid from 
threonine to alanine. This SNP is therefore a

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direct marker. Subsequent association analysis 
must determine whether this SNP affects the

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variability in the effect of leptin and, in this 
case, the indicators of fat in cattle muscle.

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The relationship between the marker and QTL 
is therefore related to their distance from

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each other and the possibility 
of crossing over between them.

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If the marker is not direct and causal, it 
may be in linkage with the causal locus QTL.

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In the figure, we can see different combinations 
of linkage disequilibrium between marker alleles

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(M1 and M2) and QTL alleles (L - worse-low and 
H - better-high performance). Depending on the

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combination on the chromosome, allele M1 may 
be advantageous in one case, according to which

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selection can be made, while allele M2 may be 
advantageous in another case. In the third case,

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no marker allele is suitable because both 
are linked to the same H allele in QTL.

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In brief, we will describe the 
principle of marker-assisted selection

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(MAS). MAS in selection programs for livestock 
allows for increased accuracy in selecting

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specific DNA variations associated with measurable 
differences in economically important traits.

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The degree of genetic improvement achieved 
through MAS can be substantially higher than

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improvement achieved through selection 
based on breeding values for traits

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with low heritability values in populations or 
traits determined post-mortem. Therefore, MAS

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has the potential to significantly increase the 
effectiveness of animal breeding for these traits.

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MAS phases. Detection phase, evaluation phase, 
and implementation phase are distinguished.

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In the detection phase, DNA polymorphisms 
are used as direct or linked markers to

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determine specific allele frequencies 
within QTL segregation populations.

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During this phase, markers associated with QTL are 
identified, and the size of allele effects and the

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location of QTL in the genome can be estimated.
In the evaluation phase, linked markers are

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tested in target populations to determine 
whether QTL segregate within the population.

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The implementation phase uses predictive 
linked markers within families, and direct

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markers are used across families to create a 
genotype database. These data are combined with

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pedigree and phenotypic information during genetic 
evaluation to predict individual genetic values.

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MAS is suitable for direct selection 
of individuals based on the genotype

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of a genetic marker/gene for simple traits 
- monogenic traits (most commonly monogenic

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diseases). For example, the CRC stress 
gene and the BLAD disease gene in cattle.

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For markers associated with quantitative 
traits, the use of MAS is limited,

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with a smaller effect - there are not as many 
described candidate genes, and there are not many

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traits with simple genetic determinism. For these 
traits, it is necessary to include whole-genome

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SNP markers in the genomic selection system.
And thank you for your attention.