this presentation is about the molecular markers as we all know the molecular markers are the DNA sequences it can be easily detected and its inheritance is easily monitored.so the main basics of the molecular markers is the polymorphic nature so it can used as molecular markers.and this will gives you the idea about AFLP, RFLP, RAPD, SNPS,ETC.
Molecular Marker and It's ApplicationsSuresh Antre
Molecular (DNA) markers are segments of DNA that can be detected through specific laboratory techniques. With the advent of marker-assisted selection (MAS), a new breeding tool is now available to make more accurate and useful selections in breeding populations.
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 observed. A genetic marker may be a short DNA sequence, such as a sequence surrounding a single base-pair change (single nucleotide polymorphism, SNP), or a long one, like minisatellites.
Molecular Marker and It's ApplicationsSuresh Antre
Molecular (DNA) markers are segments of DNA that can be detected through specific laboratory techniques. With the advent of marker-assisted selection (MAS), a new breeding tool is now available to make more accurate and useful selections in breeding populations.
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 observed. A genetic marker may be a short DNA sequence, such as a sequence surrounding a single base-pair change (single nucleotide polymorphism, SNP), or a long one, like minisatellites.
In this slide briefly describe some important note on pcr,rapd,and aflp,which helps to understand the students about this normally .
I wish for your future goal that you will shine one day inshallah .
Thank you for watching
Dr. S. MANIKANDAN, M.Sc., Ph.D
Lecturer in Botany
Thiruvalluvar University Model Constituent College,
Tittagudi 606 106, Tamil Nadu, India.
Email id: drgsmanikandan@gmail.com
In this slide briefly describe some important note on pcr,rapd,and aflp,which helps to understand the students about this normally .
I wish for your future goal that you will shine one day inshallah .
Thank you for watching
Dr. S. MANIKANDAN, M.Sc., Ph.D
Lecturer in Botany
Thiruvalluvar University Model Constituent College,
Tittagudi 606 106, Tamil Nadu, India.
Email id: drgsmanikandan@gmail.com
Marker Assisted Selection in Crop BreedingPawan Chauhan
Marker Assisted Selection is a value addition to conventional methods of Crop Breeding. It has been gaining importance in plant breeding with new generation of plant breeders and to get accurate and fast desired result from plant breeding.
I would like to share this presentation file.
Some basics information regarding to molecular plant breeding, hope this help the beginner who start working in this field.
Thanks for many original source of information (mainly from slideshare.net, IRRI, CIMMYT and any paper received from professor and some over the internet)
Molecular Breeding in Plants is an introduction to the fundamental techniques...UNIVERSITI MALAYSIA SABAH
This slide describe the process of molecular breeding in plants which involves the application of molecular markers for Marker Assisted Selection and Marker Assisted Breeding.
Molecular markers- RFLP, RAPD, AFLP, SNP etc.Cherry
Molecular markers are identifiable DNA sequences used to locate genes associated with specific traits or genetic conditions.
A molecular marker is a specific gene fragment present at a specific position called ‘locus’ (pleural loci) in the genome of a cell.
In the pool of unknown DNA or in a whole chromosome, these molecular markers help in identification of particular sequence of DNA at particular location.
Molecular marker General introduction by K. K. SAHU Sir.KAUSHAL SAHU
Introduction
Molecular marker
Characterstics of molecular marker
Types of molecular marker
. Non PCR Based
. PCR Based
RFLP
RAPD
AFLP
SSR
SNP
Conclusion
References
Molecular marker technology in studies on plant genetic diversityChanakya P
A molecular marker is a molecule contained within a sample taken from an organism (biological markers) or other matter. It can be used to reveal certain characteristics about the respective source. DNA, for example, is a molecular marker containing information about genetic disorders, genealogy and the evolutionary history of life. Specific regions of the DNA (genetic markers) are used to diagnose the autosomal recessive genetic disorder cystic fibrosis, taxonomic affinity (phylogenetics) and identity (DNA Barcoding). Further, life forms are known to shed unique chemicals, including DNA, into the environment as evidence of their presence in a particular location.Other biological markers, like proteins, are used in diagnostic tests for complex neurodegenerative disorders, such as Alzheimer's disease. Non-biological molecular markers are also used, for example, in environmental studies.
DNA Fingerprinting & its techniques by Shiv Kalia (M.Pharma in Analytical Che...Shiv Kalia
DNA fingerprinting and below mention content widely cover in this presentation
History & Introduction of DNA fingerprinting
How was the first DNA fingerprint produced?
Types of DNA Based Markers
Polymerase Chain Reaction (PCR)
PCR based Methodology of DNA fingerprinting
Electrophoresis
Utility of DNA Based Markers
Various DNA Fingerprinting Techniques Advantages & Disadvantages
Authentication of Various Ayurvedic Herbs by DNA Fingerprinting
Advantages of DNA fingerprinting in Plants
Disadvantages of DNA fingerprinting in Plants
CONCLUSION
A genetic marker is a gene or DNA sequence with a known location on a chromosome and associated with a particular gene or trait. It can be described as a variation, which may arise due to mutation or alteration in the genomic loci that can be observed. A genetic marker may be a short DNA sequence, such as a sequence surrounding a single base-pair change (single nucleotide polymorphism, SNP), or a long one, like mini & microsatellites.
RAPD markers are decamer DNA fragments.
RAPD is a type of PCR reaction.
as the name suggest it is a fast method when compared to the traditional PCR medthod.
Solid waste disposal and mangrove destructionsana sana
its describes what is waste type of wastes and solid wastes and different type of solid wastes and its disposal at last it describes mangrove destruction and its effects on environment
sand mining,food adulteration,changing crop pattern,quarringsana sana
its explain sand mining,quarrying ,changing crop pattern ,food adulteration and how it affect the environment and this each one explain each and every thing about it and the effects and some legislative method to prevent these processes
environmental movements in india and keralasana sana
this presentation describes environmental movements in india and kerala and it list number of movements in india and kerala also describes environmental and biodiversity acts
in environmental education it possess a lot of techniques that ensure the environmental education studies in higher level through higher education strategies
indutrialisation,modernisation,urbanisation and its impacts on the environmentsana sana
this slide explains the processes like urbanization modernization and industrialization and its negative impacts on environments and the solution to prevent the negative impacts
Cancer cell metabolism: special Reference to Lactate PathwayAADYARAJPANDEY1
Normal Cell Metabolism:
Cellular respiration describes the series of steps that cells use to break down sugar and other chemicals to get the energy we need to function.
Energy is stored in the bonds of glucose and when glucose is broken down, much of that energy is released.
Cell utilize energy in the form of ATP.
The first step of respiration is called glycolysis. In a series of steps, glycolysis breaks glucose into two smaller molecules - a chemical called pyruvate. A small amount of ATP is formed during this process.
Most healthy cells continue the breakdown in a second process, called the Kreb's cycle. The Kreb's cycle allows cells to “burn” the pyruvates made in glycolysis to get more ATP.
The last step in the breakdown of glucose is called oxidative phosphorylation (Ox-Phos).
It takes place in specialized cell structures called mitochondria. This process produces a large amount of ATP. Importantly, cells need oxygen to complete oxidative phosphorylation.
If a cell completes only glycolysis, only 2 molecules of ATP are made per glucose. However, if the cell completes the entire respiration process (glycolysis - Kreb's - oxidative phosphorylation), about 36 molecules of ATP are created, giving it much more energy to use.
IN CANCER CELL:
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
introduction to WARBERG PHENOMENA:
WARBURG EFFECT Usually, cancer cells are highly glycolytic (glucose addiction) and take up more glucose than do normal cells from outside.
Otto Heinrich Warburg (; 8 October 1883 – 1 August 1970) In 1931 was awarded the Nobel Prize in Physiology for his "discovery of the nature and mode of action of the respiratory enzyme.
WARNBURG EFFECT : cancer cells under aerobic (well-oxygenated) conditions to metabolize glucose to lactate (aerobic glycolysis) is known as the Warburg effect. Warburg made the observation that tumor slices consume glucose and secrete lactate at a higher rate than normal tissues.
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...Sérgio Sacani
Since volcanic activity was first discovered on Io from Voyager images in 1979, changes
on Io’s surface have been monitored from both spacecraft and ground-based telescopes.
Here, we present the highest spatial resolution images of Io ever obtained from a groundbased telescope. These images, acquired by the SHARK-VIS instrument on the Large
Binocular Telescope, show evidence of a major resurfacing event on Io’s trailing hemisphere. When compared to the most recent spacecraft images, the SHARK-VIS images
show that a plume deposit from a powerful eruption at Pillan Patera has covered part
of the long-lived Pele plume deposit. Although this type of resurfacing event may be common on Io, few have been detected due to the rarity of spacecraft visits and the previously low spatial resolution available from Earth-based telescopes. The SHARK-VIS instrument ushers in a new era of high resolution imaging of Io’s surface using adaptive
optics at visible wavelengths.
THE IMPORTANCE OF MARTIAN ATMOSPHERE SAMPLE RETURN.Sérgio Sacani
The return of a sample of near-surface atmosphere from Mars would facilitate answers to several first-order science questions surrounding the formation and evolution of the planet. One of the important aspects of terrestrial planet formation in general is the role that primary atmospheres played in influencing the chemistry and structure of the planets and their antecedents. Studies of the martian atmosphere can be used to investigate the role of a primary atmosphere in its history. Atmosphere samples would also inform our understanding of the near-surface chemistry of the planet, and ultimately the prospects for life. High-precision isotopic analyses of constituent gases are needed to address these questions, requiring that the analyses are made on returned samples rather than in situ.
What is greenhouse gasses and how many gasses are there to affect the Earth.moosaasad1975
What are greenhouse gasses how they affect the earth and its environment what is the future of the environment and earth how the weather and the climate effects.
Deep Behavioral Phenotyping in Systems Neuroscience for Functional Atlasing a...Ana Luísa Pinho
Functional Magnetic Resonance Imaging (fMRI) provides means to characterize brain activations in response to behavior. However, cognitive neuroscience has been limited to group-level effects referring to the performance of specific tasks. To obtain the functional profile of elementary cognitive mechanisms, the combination of brain responses to many tasks is required. Yet, to date, both structural atlases and parcellation-based activations do not fully account for cognitive function and still present several limitations. Further, they do not adapt overall to individual characteristics. In this talk, I will give an account of deep-behavioral phenotyping strategies, namely data-driven methods in large task-fMRI datasets, to optimize functional brain-data collection and improve inference of effects-of-interest related to mental processes. Key to this approach is the employment of fast multi-functional paradigms rich on features that can be well parametrized and, consequently, facilitate the creation of psycho-physiological constructs to be modelled with imaging data. Particular emphasis will be given to music stimuli when studying high-order cognitive mechanisms, due to their ecological nature and quality to enable complex behavior compounded by discrete entities. I will also discuss how deep-behavioral phenotyping and individualized models applied to neuroimaging data can better account for the subject-specific organization of domain-general cognitive systems in the human brain. Finally, the accumulation of functional brain signatures brings the possibility to clarify relationships among tasks and create a univocal link between brain systems and mental functions through: (1) the development of ontologies proposing an organization of cognitive processes; and (2) brain-network taxonomies describing functional specialization. To this end, tools to improve commensurability in cognitive science are necessary, such as public repositories, ontology-based platforms and automated meta-analysis tools. I will thus discuss some brain-atlasing resources currently under development, and their applicability in cognitive as well as clinical neuroscience.
Richard's aventures in two entangled wonderlandsRichard Gill
Since the loophole-free Bell experiments of 2020 and the Nobel prizes in physics of 2022, critics of Bell's work have retreated to the fortress of super-determinism. Now, super-determinism is a derogatory word - it just means "determinism". Palmer, Hance and Hossenfelder argue that quantum mechanics and determinism are not incompatible, using a sophisticated mathematical construction based on a subtle thinning of allowed states and measurements in quantum mechanics, such that what is left appears to make Bell's argument fail, without altering the empirical predictions of quantum mechanics. I think however that it is a smoke screen, and the slogan "lost in math" comes to my mind. I will discuss some other recent disproofs of Bell's theorem using the language of causality based on causal graphs. Causal thinking is also central to law and justice. I will mention surprising connections to my work on serial killer nurse cases, in particular the Dutch case of Lucia de Berk and the current UK case of Lucy Letby.
Introduction:
RNA interference (RNAi) or Post-Transcriptional Gene Silencing (PTGS) is an important biological process for modulating eukaryotic gene expression.
It is highly conserved process of posttranscriptional gene silencing by which double stranded RNA (dsRNA) causes sequence-specific degradation of mRNA sequences.
dsRNA-induced gene silencing (RNAi) is reported in a wide range of eukaryotes ranging from worms, insects, mammals and plants.
This process mediates resistance to both endogenous parasitic and exogenous pathogenic nucleic acids, and regulates the expression of protein-coding genes.
What are small ncRNAs?
micro RNA (miRNA)
short interfering RNA (siRNA)
Properties of small non-coding RNA:
Involved in silencing mRNA transcripts.
Called “small” because they are usually only about 21-24 nucleotides long.
Synthesized by first cutting up longer precursor sequences (like the 61nt one that Lee discovered).
Silence an mRNA by base pairing with some sequence on the mRNA.
Discovery of siRNA?
The first small RNA:
In 1993 Rosalind Lee (Victor Ambros lab) was studying a non- coding gene in C. elegans, lin-4, that was involved in silencing of another gene, lin-14, at the appropriate time in the
development of the worm C. elegans.
Two small transcripts of lin-4 (22nt and 61nt) were found to be complementary to a sequence in the 3' UTR of lin-14.
Because lin-4 encoded no protein, she deduced that it must be these transcripts that are causing the silencing by RNA-RNA interactions.
Types of RNAi ( non coding RNA)
MiRNA
Length (23-25 nt)
Trans acting
Binds with target MRNA in mismatch
Translation inhibition
Si RNA
Length 21 nt.
Cis acting
Bind with target Mrna in perfect complementary sequence
Piwi-RNA
Length ; 25 to 36 nt.
Expressed in Germ Cells
Regulates trnasposomes activity
MECHANISM OF RNAI:
First the double-stranded RNA teams up with a protein complex named Dicer, which cuts the long RNA into short pieces.
Then another protein complex called RISC (RNA-induced silencing complex) discards one of the two RNA strands.
The RISC-docked, single-stranded RNA then pairs with the homologous mRNA and destroys it.
THE RISC COMPLEX:
RISC is large(>500kD) RNA multi- protein Binding complex which triggers MRNA degradation in response to MRNA
Unwinding of double stranded Si RNA by ATP independent Helicase
Active component of RISC is Ago proteins( ENDONUCLEASE) which cleave target MRNA.
DICER: endonuclease (RNase Family III)
Argonaute: Central Component of the RNA-Induced Silencing Complex (RISC)
One strand of the dsRNA produced by Dicer is retained in the RISC complex in association with Argonaute
ARGONAUTE PROTEIN :
1.PAZ(PIWI/Argonaute/ Zwille)- Recognition of target MRNA
2.PIWI (p-element induced wimpy Testis)- breaks Phosphodiester bond of mRNA.)RNAse H activity.
MiRNA:
The Double-stranded RNAs are naturally produced in eukaryotic cells during development, and they have a key role in regulating gene expression .
2. •A DNA sequence that is readily detected and whose inheritance can be easily monitored.
•The uses of molecular markers are based on the naturally occurring polymorphism.
•A marker is a gene of known function and location, that allow the studying of the
inheritance of the gene.
•A marker must be a polymorphic i.e., it must exist in different forms so that chromosomes
carrying mutant gene can be distinguished from the chromosome with the normal gene
by a marker.
• NB: polymorphism involves existence of different forms of same gene in plants or
population of plants.
Examples: RFLP,RAPD,AFLP, VNTR,SSR,SNP etc…
3. They may be due to:
• Base pair changes.
• Rearrangements (translocation or
inversion).
• Insertions or deletions.
• Variation in the number of tandem
repeats.
THE MARKERS REVEALING VARIATION AT DNA LEVEL
ARE REFERRED TO AS THE MOLECULAR MARKERS.
4. Protein markers such as alloenzymes
DNA markers such as:
MITOCHONDRIAL DNA MARKERS
NUCLEAR DNA MARKERS ( RAPD ,
VNTR SUCH AS MICRO & MINI SATELLITES)
5. A MOLECULAR MARKER MUST POSSES THE FOLLOWING
DESIRABLE PROPERTIES
It must be polymorphic ( different structures in
different individuals of a species ) so that diversity
must be measured
It should be evenly distributed throughout the
genome
It should distinguishes the homozygotes and
heterozygotes.
6. In the organism ,the polymorphism can be distinguished
at three levels
1. Morphological or phenotypic; i.e., The morphological marker
corresponds to a quantitative trait scored visually
2. Biochemical marker due to difference in protein
e.g.; Isozymes
3. Molecular marker-DNA sequences among which the differences
can be detected and monitored in the subsequent generation
7. Advantages of Molecular Markers
Ubiquitous.
Stably inherited.
Multiple alleles for each marker.
Devoid of pleiotropic effects.
Detectable in all tissues, at all ages.
Long shelf life of the DNA samples.
8. RFLP – (RESTRICTION FRAGMENT LENGTH POLYMORPHISM)
•Genetic markers
resulting from the
variation or change in
the length of defined
DNA fragments
produced by digestion
of the DNA sample with
restriction
endonucleases
9. RFLP is a difference in homologous DNA sequences that can be detected by
presence of fragment of different lengths after digestion of the DNA samples
with specific restriction endonucleases.
When the genomic DNA of many individuals of one species are separately
cleaved by restriction enzymes ,passed through electrophoresis.
Then it blotted on nitrocellulose membrane and probed with a radio labelled
DNA.
There arises polymorphism in hybridisation pattern of digested DNAs
This shows differences in sequences between two individual. These unique,
common and repeated DNA sequences found in several individuals are
termed as RFLPS.
After digestion with specific enzymes, variations obtained in one DNA
fragment with that enzyme is referred to as one RFLP.
10. The RFLPs are detected by using the unique sequence of DNA called genomic probe,
RFLP map constructed as follows
Large molecular weight genomic DNA is isolated from several strains or related species
These DNAs are then digested with a selected restriction enzyme. The fragments in these
digest are separated through electrophoresis
The resulting gel lines are transferred to a suitable solid support and exposed to e radio
labelled probe under conditions favouring hybridisation
The free probes are removed and finally the fragments to which the probe has hybridised
are detected by autoradiography
An RFLP occurs when the length of a detected fragments varies between individuals
Each fragment length is considered an allele ,and can be used in genetic analysis
11. Microbial cell or plant / animal tissue
Genomic DNA
DNA fragments separated on gel
DNA on nitrocellulose membrane
Southern hybridisation
Isolate DNA
Digest DNA with restriction enzymes
Separate on agarose gel
Southern blotting
Incubate with suitable genome cloned probe
autoradiography
Positive band shows RFLP PATTERN
13. •High reproducibility
•Show co-dominant alleles
•Detect coupling phase of DNA
•Reliable marker in linkage and breeding analysis
•Easily determine a linked trait present in both homozygous and heterozygous
ADVANTAGES
14. •Require large quantities of high molecular weight DNA.
•Expensive process
•Time consuming
•Labor intensive
DISADVANTAGES
15. CONSIDERATIONS FOR USE OF RFLPS
-Relatively slow process
-Use of radioisotopes has limited RFLP use to
certified laboratories (but non-radioactive labeling
systems are now in wide use)
- Co-dominant markers; often species-specific
- Need high quality DNA
- Need to develop polymorphic probes - expensive
16.
17. AFLP is based on a selectively amplifying a subset of restriction fragments from a
complex mixture of DNA fragments obtained after digestion of genomic DNA with
restriction endonucleases.
Polymorphisms are detected from differences in the length of the amplified
fragments by polyacrylamide gel electrophoresis (PAGE) .
The technique involves four steps: (1) restriction of DNA and
ligation of oligonucleotide adapters (2) preselective amplification (3)
selective amplification (4) gel analysis of amplified fragments.
AFLP involves the restriction of genomic DNA, followed by ligation of adaptors
complementary to the restriction sites and selective PCR amplification of a subset of the
adapted restriction fragments. These fragments are viewed on denaturing
polyacrylamide gels either through autoradiographic or fluorescence methodologies .
AFLP(AMPLIFIED FRAGMENT LENGTH POLYMORPHISM)
18. High genomic abundance.
Considerable reproducibility.
AFLPs can be analyzed on automatic sequencers.
The generation of many informative bands per reaction.
Capability to amplify between 50 and 100 fragments at
one time.
Higher resolution and sensitivity.
ADVANTAGES
19. • Need for purified, high molecular
weight DNA.
• The major disadvantage of AFLP
markers is that these are dominant
markers.
•Abundance of data.
DISADVANTAGES
20. •AFLPs can be applied in studies involving genetic identity, parentage and
identification of clones and cultivars.
•phylogenetic studies of closely related species.
• AFLP markers have successfully been used for analyzing genetic diversity in some
other plant species such as peanut.
•This technique is useful for breeders to accelerate plant improvement.
•AFLP markers are useful in genetic studies, such as biodiversity evaluation,
analysis of germplasm collections, genotyping of individuals and genetic distance
analyses.
APPLICATIONS
21. RAPD ( RANDOM AMPLIFIED POLYMORPHIC DNA )
•It is a PCR based technology.
•In 1991 Welsh and Maclelland developed this technique.
•This procedure detects nucleotide sequence polymorphism in DNA.
•It is used to analyze genetic diversity of an individual by random primers.
• RAPD generates a type of genetic finger print that can be used to identify individuals
• If we have some information about a set of fragments randomly distributed through out the
genome ,we may synthesize a single short oligonucleotide (10bp) and use both ends of DNA
segments before its amplification
• The DNA amplification product is generate from a region flanked by a pair of 10bp primary
sites, consequently ,random sample of DNA markers is obtained which is called RAPD
• Unlike the traditional PCR analysis ,RAPD does not require any specific knowledge of the DNA
sequences of the target organisms.
• Genomic DNA of two individuals produce different RAPDs.
22. •A specific DNA segment shows DNA polymorphism. Hence it can be used as a
genetic marker
• Many random oligonucleotide primers are designed by using different
combination of nucleotides.
• This makes the method popular for comparing the DNA of biological systems that
have not had the attention of the scientific community
• It has some limitation in the use of degraded DNA samples .
• It’s resolving power is much lower than targeted ,species specific DNA
comparison methods ,such as short tandem repeats .
• In recent years ,RAPD has been used to characterize ,and trace,
the phylogeny of diverse plant and animal species.
23.
24. Cell /tissue of an individual
Double stranded genomic DNA
Single stranded DNA
Primers annealed to template DNA
Complementary strand is synthesized
Isolate DNA
-Taq polymerases ,primer, dNTPs
- Denature DNA @ 94 c° for 1 minute
- keep in tubes of PCR thermocycler
Annealing of primers ( 36° c for 2 min)
DNA is synthesized at 72° c for 1.5 min
Separate amplified products by gene electrophoresis
35 – 45 cycles
25. •Quick and easy to assay.
•Low quantities of template DNA required.
•Dominant markers.
•In expensive.
•Do not require any specific knowledge of the target.
• No species specific probes are required for different species .
• Crude DNA preparation may be used for analysis of whole genome
• It does not require blotting or hybridization
26. •Low reproducibility
•Highly sensitive and complicated procedure.
•PCR cycling conditions greatly influence the out come.
•Mismatches between primer and template may result in total absence of
PCR product.
• RAPD markers are dominant so it is cause problems whether the DNA
segment is amplified from locus that heterozygous ,homozygous.
27. •Gene mapping
•DNA amplification finger printing
•Study of closely related species
• RAPD technique include Arbitrarily Primed Polymerase Chain Reaction (AP-
PCR).
• Certain genetic markers may be tagged at specific regions in the genome
28. •The term microsatellites was coined by Litt & Lutty (1989)and it also known as Simple
Sequence Repeats (SSRs), are sections of DNA.
•Microsatellite markers, developed from genomic libraries, can belong to either the
transcribed region or the non transcribed region of the genome.
• Microsatellite sequences are especially suited to distinguish closely related genotypes;
because of their high degree of variability, they are, therefore, favoured in population
studies .
• Microsatellite polymorphism can be detected by Southern hybridization or PCR.
•If nucleotide sequences in the flanking regions of the microsatellite are known, specific
primers can be designed to amplify the microsatellite by PCR.
•microsatellite may be identified by screening sequence databases,
polymorphism can detected by gel electrophoresis
SSR (SIMPLE SEQUENCE REPEAT) OR MICROSATELLITES
29. Microsatellites – Properties
Co-dominant
Inherit in Mendelian Fashion
Polymorphic loci with allele number as high as 14 – 15 per
locus
Mostly reported from non-coding region, hence can be
independent of selection
Flanking region is highly conserved in related species
Can be obtained from small amounts of tissues [STR
analysis can be done on less than one billionth of a gram
(a nanogram) of DNA (as in a single flake of dandruff)]
PAGE separation; silver staining/automated genotyping
Abundant in the eukaryote genome (~103 to 105 loci
dispersed at 7 to 10100 kilobase pair (kb) intervals)
30. Also called as STR, SSR.
Tandemly repeated DNA sequences with the repeat/size
of 1 – 6 bases repeated several times
Highly polymorphic; can be analyzed with the help of
PCR
Individual alleles at a locus differ in number of tandem
repeats of unit sequence owing to gain of loss of one or
more repeats and they can be differentiated by
electrophoresis according to their size
Powerful DNA markers for quantifying genetic variations
within & between populations of a species
31. Short Tandem Repeats (STRs)
the repeat region is variable between samples while the
flanking regions where PCR primers bind are constant
7 repeats
8 repeats
Homozygote = both alleles are the same length
Heterozygote = alleles differ and can be resolved
from one another
32. •The microsatellite, or short sequence repeat (SSR), is
a powerful genetic marker, useful in many areas of
fish genetics and breeding.
•Polymorphic microsatellite loci have been frequently
applied to the analysis of genetic diversity, population
genetic structure, and genomic mapping.
•These co-dominant markers have also been applied
to the classification and systematics, parentage
identification, germplasm conservation, and breeding
programme of food fish.
33. • Because the technique is PCR-based, only low quantities of template DNA
(10–100 mg per reaction) are required.
•The strengths of microsatellites include the co-dominance of alleles, their
high genomic abundance
• the reproducibility of microsatellites is high and analyses do not require high
quality DNA
34. • Main drawbacks of microsatellites is that high development costs
• Errors in genotype scoring
•Difficulty in interpretation
• Previous genetic information is needed
• Huge upfront work required
• Problems associated with PCR of microsatellites
35.
36. FORENSICS
•Microsatellite loci are widely used for forensic identification and relatedness testing and
are predominant genetic marker in this area of application
•In forensic identification cases ,the goal is typically to link a suspect with a sample of
blood semen, or hair taken from a crime
•Another application involves linking DNA samples with relative of a missing person,
because the length of microsatellite may vary from one person to next .(finger printing)
DIAGNOSIS AND IDENTIFICATION OF HUMAN DISEASES
• Microsatellites changes in length early in the development of some cancers , they are
useful markers for early cancer detection
• It is useful in linkage studies so it help to locate genes responsible for various genetic
disorders
37. POPULATION STUDIES
• By looking at the variation of microsatellites in populations,
inferences can be made about population structures and differences ,
genetic drift, genetic bottlenecks and even date of a last common
ancestor.
CONSERVATION BIOLOGY
• Used to detect sudden changes in population
• Effect of population fragmentation
• Interaction of different population
• useful in identification of new and incipient populations
38. MINISATELLITE DNAs
•These are usually 0.2 to 2 kb long and consists of tandem repeats of 9
to 40 bp sequences
• In the case of humans ,minisatellite DNAs are concentrated in the
terminal regions of the chromosomes
• Therefore they do not contribute a good marker system for human
genome ,despite the great degree of polymorphism exhibited by them
• They are useful in the finger printing
39. VNTR ( VARIABLE NUMBER OF TANDEM REPEATS)
•Repetitive DNA composed of copies of short sequences, involved in generation of
polymorphic loci that are useful in genetic fingerprinting , also known as hyper variable
regions.
• A variable number of tandem repeat is a location in a genome where a short
nucleotide sequence is organized as a tandem repeat.
• Tandem repeats occur in DNA when a pattern of 2 or more nucleotide is repeated and
the repetitions are directly adjacent to each other.
• e.g.; A-T-T-C-G-A-T-T-C-G-A-T-T-C-G In which the sequence A-T-T-C-G is repeated 3
times
• These can be found on many chromosomes ,and often show variations in length
between individuals. each variant act as an inherited allele, allowing them to be used
for personal or parental identification
•Their analysis is useful in genetics and biology research , forensic, and DNA finger
printing
40. • VNTRs are dispersed throughout the genome and are made up of a variable number of
end to end duplications of identical or almost identical sequences of 12 -80 bp each
• polymorphism in VNTRs is usually associated with the number of repeats.
• These variations constitute the allele of VNTR loci, polymorphism also arises due to a
variation in the distribution pattern of repeated sequences in the different alleles.
• VNTRs were in important source of RFLP genetic markers used in linkage analysis of
genome
• VNTRs have become essential to forensic crime investigation, via fingerprinting
•When removed from surrounding DNA by PCR or RFLP methods, and their size determined
by gel electrophoresis or southern blotting ,they produce a pattern of bands unique to
each individual.
• VNTRs analysis is also being used to study genetic diversity and breeding patterns in
populations of wild or domesticated animals
41. SNPs ( single nucleotide polymorphism)
GCTGTATGACTAGAAGATCG
AT
GCTGTATGACGAGAAGATCG
AT•IT IS VARIATION IN A SINGLE NUCLEOTIDE WHICH MAY OCCUR AT SOME SPECIFIC POSITION
IN THE GENOME WHERE EACH VARIATION IS PRESENT TO SOME APPRECIABLE DEGREE
WITH IN A POPULATION.
OR
•A SNP IS DEFINED AS A SINGLE BASE CHANGE IN A DNA SEQUENCE THAT OCCURS IN A
SIGNIFICANT PROPORTION (MORE THAN 1 PERCENT) OF A LARGE POPULATION.
42. •In human beings, 99.9 percent bases are same.
•Remaining 0.1 percent makes a person unique.
•Different attributes / characteristics / traits
•how a person looks,
•diseases he or she develops.
•These variations can be:
•Harmless (change in phenotype)
•Harmful (diabetes, cancer, heart disease, Huntington's disease,
and hemophilia )
•Latent (variations found in coding and regulatory regions, are not
harmful on their own, and the change in each gene only becomes
apparent under certain conditions e.g. susceptibility to lung
cancer)
43. ►SNPs are found in
coding and (mostly) noncoding regions.
►Occur with a very high frequency
about 1 in 1000 bases to 1 in 100 to 300 bases.
►The abundance of SNPs and the ease with which they can be
measured make these genetic variations significant.
►SNPs close to particular gene acts as a marker for that gene.
►SNPs in coding regions may alter the protein structure made by
that coding region.
SNP FACTS
44. •Single-nucleotide polymorphisms may fall within coding sequences of genes, non-
coding regions of genes, or in the intergenic regions (regions between genes).
•SNPs within a coding sequence do not necessarily change the amino acid sequence
of the protein that is produced, due to degeneracy of the genetic code.
•SNPs in the coding region are of two types,
• synonymous
• nonsynonymous
• Synonymous SNPs do not affect the protein sequence
•nonsynonymous change the amino acid sequence of protein. The nonsynonymous
SNPs are of two types: missense and nonsense.
45. IN DISEASE DIAGNOSIS
IN FINDING PREDISPOSITION TO DISEASES
IN DRUG DISCOVERY & DEVELOPMENT
IN DRUG RESPONSES
INVESTIGATION OF MIGRATION PATTERNS
ALL THESE ASPECT WILL HELP TO LOOK FOR MEDICATION &
DIAGNOSIS AT INDIVIDUAL LEVEL
SIGNIFICANCE OF SNPs
46. 1. SNPs ARE THE MOST FREQUENT FORM OF DNA
VARIATIONS
2. THEY ARE THE DISEASE CAUSING MUTATIONS IN
MANY GENES
3. THEY ARE ABUNDANT & HAVE SLOW MUTATION RATES
4. EASY TO SCORE
5. MAY WORK AS THE NEXT GENERATION OF GENETIC
MARKERS
47. •SNPs are used for identification and forensics
•SNPs are used for mapping and genome-wide
association studies of complex diseases
•SNPs are used for estimating predisposition to
disease
•SNPs are used for immigration & citizenship in the UK
•SNPs are used to predict specific genetic traits
•SNPs are used for classifying patients in clinical trials