seminar, cvas bikaner

1. DEPARTMENT OF
ANIMAL BREEDING & GENETICS
(MAS)
MARKER ASSISTED SELECTION
SUBMITTED BY :
SUNIL KUMAR BHAKAR

2. OVERVIEW
 DEFINITION
 MARKER TYPES
 PROPERTY OF IDEAL MARKER FOR MAS
 POSITIVE AND NEGATIVE SELECTABLE MARKERS
 GENE VS MARKER
 RELATION B/W MARKER AND ASSOCIATED GENE
 STEPS FOR MAS
 FACTOR FOR SUCCESS
 MAS WORK BEST FOR TRAITS
 HIGH THROUGHPUT GENOTYPING TECHNIQUES
 CONCLUSION

3. What is MAS
 Marker assisted selection or marker aided
selection (MAS) is a process whereby a marker
(morphological, biochemical molecular etc.) is used for
indirect selection of a genetic determinant or determinants of a
trait of interest (e.g. productivity, disease resistance, abiotic
stress tolerance etc..).
 This process is used in plant and animal breeding.

4. POSITIVE AND NEGATIVE
SELECTABLE MARKER
 Positive selectable markers are selectable markers that
confers selective advantage to its host organism. An
example would be antibiotics resistance, which allows the
host organism to survive antibiotics selection.
 Negative selectable markers are selectable markers that
would eliminate its host organism upon selection. An
example would be thymidine kinase, which would make the
host sensitive to ganciclovir selection.
MARKER TYPES

5. Biological markers
Different pathogen races or insect biotypes based on
host pathogen or host parasite interaction can be used
as a marker since the genetic constitution of an
organism can affect its susceptibility to pathogens or
parasites.

6. Morphological markers
 Animals are selected based on appearance.
 Disadvantage: lack of polymorphism

7. Biochemical markers
Animal are selected on the basis of biochemical properties.
Eg. Hb, AMYLASE, BLOOD GROUPS ETC.
Disadvantage:
Sex limited
Age dependent
Influenced by environment
It covers less than 10% of genome

8. Cytological markers
The chromosomal banding produced by different stains; for
example, G banding.
structural and numerical variations are identified.
Structural- Deletions, Insertions etc.
Numerical- Trisomy , Monosomy , Nullysomy
Disadvantage : low polymorphism

9. DNA-based / molecular
 To avoid problems specific to morphological markers, the DNA-based markers have
been developed.
 A unique (DNA sequence), occurring in proximity to the gene or locus of interest, can
be identified by a range of molecular techniques.
 Three common technologies used as molecular markers are: RFLP, SSR
(microsatellites) and SNP .
 ADVANTASES
1. Highly polymorphic.
2. Simple inheritance.
3. Abundantly occur throughout the genome.
4. Easy and fast to detect,
5. Minimum pleiotropic effect.
6. Detection is not dependent on the
developmental stage of the organism.

10. Important Properties of
Ideal Markers for MAS
 Easy recognition of all possible phenotypes(homo-
and heterozygote's) from all different alleles.
 Demonstrates measurable differences in expression
between trait types and/or gene of interest alleles, early in
the development of the organism.
 Low or null interaction among the markers allowing the use
of many at the same time in a segregating population.
 Abundant in number.
 Polymorphic.

11. Gene Vs Marker
 The gene of interest is directly related with production of
protein(s) that produce certain phenotypes whereas
markers not influence the trait of interest but are genetically
linked .
 In many traits, genes are discovered and can be directly
assayed for their presence with a high level of confidence.
However, if a gene is not isolated, markers help is taken to
tag a gene of interest. In such case there may be some
inaccurate (even false) positive results due to
recombination between the marker of interest and gene (or
QTL). A perfect marker would elicit no false positive results.

12. Relationships b/w
markers and respective gene
Three kinds of relationships between the markers and
respective genes could be distinguished :
(1) The molecular marker located within the gene of interest,
which is the most favourable situation for MAS and in this
case, it could be ideally referred to as gene-assisted selection.
Such markers are called as direct marker.
(2) The marker in linkage disequilibrium (LD) with the gene of
interest throughout the population. LD is the tendency of certain
combination of alleles to be inherited together. Population-wide
LD can be found when markers and genes
of interest are physically close to each other. Selection using
these markers can be called as LD-MAS.
(3) The marker in linkage equilibrium (LE) with the gene of interest
throughout the population, which is the most difficult and
challenging situation for applying MAS.

13. Cont…
 Direct markers are preferred for effective implementation of
marker-assisted selection, followed by LD and LE markers,
the latter requiring within-family analysis and selection.
 Ease of application and potential for extra-genetic gain is
greatest for direct markers, followed by LD markers, but is
antagonistic to ease of detection, which is greatest for LE
markers.

14. Steps for MAS
 Generally the first step is to map the gene or quantitative
trait locus (QTL) of interest by using different techniques
and then use 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.

15. CONT….
 Three common technologies used as molecular markers are:
restriction fragment length polymorphisms, simple sequence
repeats, and single nucleotide polymorphisms.
 Genetic variants often differ from each other by the sequence
of a single nucleotide so single nucleotide polymorphisms
SNPs (pronounced “snips”),is commonly the basis of
genotyping tests.
 It is therefore possible to genotype an animal using a DNA-
based genotyping test and select individuals carrying the
preferred genetic variant.

16. Factor for success of MAS
In general , the success of MAS depends on following factor :-
(i) Availability of genetic map with an adequate number of
uniformly-spaced polymorphic markers to accurately locate
desired QTLs or major gene(s).
(ii) Close linkage between the QTL or a major gene
of interest and adjacent markers.
(iii) Ideally markers should be <5 cM from a gene or QTL
Marker A
QTL5 cM
RELIABILITY FOR
SELECTION
Using marker A only:
1 – rA = ~95%
Marker A
QTL
Marker B
5 cM 5 cM
Using markers A and B:
1 - 2 rArB = ~99.5%

17. Cont…
Note- Using a pair of flanking markers can
greatly improve reliability but increases time
and cost.
(iv) Adequate recombination between the markers
and rest of the genome.
(v) Ability to analyze a larger number of Animals
in a time and cost effective manner.

18. Fundament Advantages of MAS
1. Greater efficiency.
2. Accelerated line development in breeding programs.
For example, time and labour savings may arise from the
substitution of difficult or time-consuming field trials (that
need to be conducted at particular times of year or at
specific locations, or are technically complicated) with DNA
marker tests.
3 .Selection based on DNA markers may be more reliable
due to the influence of environmental factors on field trials.

19. CONT…
4. Another benefit from using MAS is that the total number of
lines that need to be tested may be reduced considerably.
5. Since many lines can be discarded after MAS at an early
generation, this permits a more effective breeding design.
6. Greater efficiency of target trait selection may enable
certain traits to be fast-tracked, thus specific genotypes
can be easily identified and selected.

20. MAS work best for traits
 1. Low h2
 2. Are difficult or expensive to measure.
e.g. disease resistance.
 3. Can not be measured untill after selection has occurred
e.g. Carcass data.
 4 .Are currently not selected for due to lack of available
phenotypic data.
e.g.(tenderness).

21. Traits most likely to benefit for MAS
(descending order ).
1. Disease resistance .
2. Carcass quality and palatability attributes.
3. Fertility and reproductive efficiency.
4. Maintenance requirements.
5. Carcass quantity and yields.
6. Milk production and maternal ability.
7. Growth performance.

22. 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 of example of such automation is using DNA
isolation robots, capillary electrophoresis and pipetting robots.
 One of recent example of
capillary system is Applied
Bio systems 3130 Genetic
Analyser.

23. CONCLUSIONS
 In context of MAS, markers can be effectively utilized for two
basic purposes :-
(i) tracing favourable allele(s) (dominant or recessive) across
generations.
(ii) Identifying the most suitable individual(s) among the
segregating progeny, based on allelic composition across a part
or the entire genome.
 It is important to combine DNA results (which look at single
genes) with other criteria, such as EPDs (which look at
numerous genes) and the animal’s actual phenotype for the trait
(if available), to ensure that selection is distributed over all the
genes that contribute towards the trait of interest.

24. CONT….
 Don't ignore animals that have good EPDs for a given trait and
yet are not carrying the favorable form of a gene for that trait.
These animals are likely to be a source of good alleles for the
many other genes that contribute towards that trait.
 Ideally the information from genetic tests should be integrated
into a genetic evaluation system that weighs all the information
about an animal. Combining information from both EPDs and
genetic tests into a selection decision will be superior to
selection on either EPDs or markers alone.