Epigenetics and it's relevance in crop improvementShamlyGupta
Epigenetics means ‘above’ or ‘on top of genetics’
A study of the changes in gene expression that are mitotically and/or meiotically heritable and do not involve a change in the DNA sequence
Gene-regulatory information that is not expressed in DNA sequences but transmitted from one generation (of cells or organisms) to the next
Coined by embryologist C. H. Waddington in 1942.
A concise and well fabricated presentation the current techniques used for plant genome editing including CRISPER/cas9 system, TALENS, TELES, ZINC FINGER NUCLEASES(ZFN), HEJ (homologous endjoing) and many other high throughout techniques along references.
A new era of genomics for plant science research has opened due the complete genome sequencing projects of Arabidopsis thaliana and rice. The sequence information available in public database has highlighted the need to develop genome scale reverse genetic strategies for functional analysis (Till et al., 2003). As most of the phenotypes are obscure, the forward genetics can hardly meet the demand of a high throughput and large-scale survey of gene functions. Targeting Induced Local Lesions in Genome TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identity point mutations in regions of interest (McCallum et al., 2000). This strategy works with a mismatch-specific endonuclease to detect induced or natural DNA polymorphisms in genes of interest. A newly developed general reverse genetic strategy helps to locate an allelic series of induced point mutations in genes of interest. It allows the rapid and inexpensive detection of induced point mutations in populations of physically or chemically mutagenized individuals. To create an induced population with the use of physical/chemical mutagens is the first prerequisite for TILLING approach. Most of the plant species are compatible with this technique due to their self-fertilized nature and the seeds produced by these plants can be stored for long periods of time (Borevitz et al., 2003). The seeds are treated with mutagens and raised to harvest M1 plants, which are consequently, self-fertilized to raise the M2 population. DNA extracted from M2 plants is used in mutational screening (Colbert et al., 2001). To avoid mixing of the same mutation only one M2 plant from each M1 is used for DNA extraction (Till et al., 2007). The M3 seeds produce by selfing the M2 progeny can be well preserved for long term storage. Ethyl methane sulfonate (EMS) has been extensively used as a chemical mutagen in TILLING studies in plants to generate mutant populations, although other mutagens can be effective. EMS produces transitional mutations (G/C, A/T) by alkylating G residues which pairs with T instead of the conservative base pairing with C (Nagy et al., 2003). It is a constructive approach for users to attempt a range of chemical mutagens to assess the lethality and sterility on germinal tissue before creating large mutant populations.
Epigenetics and it's relevance in crop improvementShamlyGupta
Epigenetics means ‘above’ or ‘on top of genetics’
A study of the changes in gene expression that are mitotically and/or meiotically heritable and do not involve a change in the DNA sequence
Gene-regulatory information that is not expressed in DNA sequences but transmitted from one generation (of cells or organisms) to the next
Coined by embryologist C. H. Waddington in 1942.
A concise and well fabricated presentation the current techniques used for plant genome editing including CRISPER/cas9 system, TALENS, TELES, ZINC FINGER NUCLEASES(ZFN), HEJ (homologous endjoing) and many other high throughout techniques along references.
A new era of genomics for plant science research has opened due the complete genome sequencing projects of Arabidopsis thaliana and rice. The sequence information available in public database has highlighted the need to develop genome scale reverse genetic strategies for functional analysis (Till et al., 2003). As most of the phenotypes are obscure, the forward genetics can hardly meet the demand of a high throughput and large-scale survey of gene functions. Targeting Induced Local Lesions in Genome TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identity point mutations in regions of interest (McCallum et al., 2000). This strategy works with a mismatch-specific endonuclease to detect induced or natural DNA polymorphisms in genes of interest. A newly developed general reverse genetic strategy helps to locate an allelic series of induced point mutations in genes of interest. It allows the rapid and inexpensive detection of induced point mutations in populations of physically or chemically mutagenized individuals. To create an induced population with the use of physical/chemical mutagens is the first prerequisite for TILLING approach. Most of the plant species are compatible with this technique due to their self-fertilized nature and the seeds produced by these plants can be stored for long periods of time (Borevitz et al., 2003). The seeds are treated with mutagens and raised to harvest M1 plants, which are consequently, self-fertilized to raise the M2 population. DNA extracted from M2 plants is used in mutational screening (Colbert et al., 2001). To avoid mixing of the same mutation only one M2 plant from each M1 is used for DNA extraction (Till et al., 2007). The M3 seeds produce by selfing the M2 progeny can be well preserved for long term storage. Ethyl methane sulfonate (EMS) has been extensively used as a chemical mutagen in TILLING studies in plants to generate mutant populations, although other mutagens can be effective. EMS produces transitional mutations (G/C, A/T) by alkylating G residues which pairs with T instead of the conservative base pairing with C (Nagy et al., 2003). It is a constructive approach for users to attempt a range of chemical mutagens to assess the lethality and sterility on germinal tissue before creating large mutant populations.
Plant epigenetic memory in plant growth behavior and stress response. Sally M...CIAT
Speaker: Sally Mackenzie, Lloyd and Dottie Huck Chair for Functional Genomics, Department of Biology, Pennsylvania State University. Fellow in the American Society of Plant Biologists and the American Association for the Advancement of Science (AAAS).
Event: Robert D. Havener Seminar on “Innovations for Crop Productivity”.
http://ciat.cgiar.org/event/robert-d-havener-seminar-on-innovations-for-crop-productivity/
Access to large-scale omics datasets i.e. genomics, transcriptomics, proteomics, metabolomics, phenomics, etc. has revolutionized biology and led to the emergence of systems approaches to advance our understanding of biological processes. With decreasing time and cost to generate these datasets, omics data integration has created both exciting opportunities and immense challenges for biologists, computational biologists, biostatisticians and biomathematicians. Genomics, transcriptomics, proteomics, and metabolomics together they help to bring out the best of characters in plants.
Association mapping, also known as "linkage disequilibrium mapping", is a method of mapping quantitative trait loci (QTLs) that takes advantage of linkage disequilibrium to link phenotypes to genotypes.Varioius strategey involved in association mapping is discussed in this presentation
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.
Targeted Induced Local Lesions IN Genome. Mutations (Single base pair substitution) are created by traditionally used chemical mutagens. Identify SNPs and / or INDELS in a gene / genes of interest from a mutagenized population.
Majority of agronomic traits are quantitative and are controlled polygenetically.Instead of producing transgenic plants through single gene transfer many researchers are attempting on multigene engineering. The simultaneous transfer of multiple genes in to plants will enable us to produce plants with more desirable characters. Engineering of genes coding for complete metabolic pathways, bacterial operons or biopharmaceuticals that require an assembly of complex multisubunit proteins etc are some of the successful examples of multigene engineering.
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)
Current Status of TILLING and EcoTILLING:
TILLING and EcoTILLING technique have been adapted in diverse species including rice, maize, Lotus, poplar, Arabidopsis, wheat, barley, potato, tomato, sunflower, common bean, Field Mustard, clover, melon, pea, peanut, sorghum, rapeseed, soybean, melon, poplar, sugarcane, brassica and other for the purpose of gene detection, functional genomics, polymorphism assessment, plant breeding as described in case study part.
Ecotilling:
EcoTILLING is similar to TILLING, except that its objective is to identify natural genetic variation as opposed to induced mutations.
Many species are not amenable to chemical mutagenesis; therefore, EcoTILLING can aid in the discovery of natural variants and their putative gene function
This approach allows one to rapidly screen through many samples with a gene of interest to identify naturally occurring SNPs and / or small INs/DELS
iTILLING:
A new approach to the TILLING method that reduces costs and the time necessary to carry out mutation screening was developed for Arabidopsis and it is called iTILLING, individualized TILLING
Plant epigenetic memory in plant growth behavior and stress response. Sally M...CIAT
Speaker: Sally Mackenzie, Lloyd and Dottie Huck Chair for Functional Genomics, Department of Biology, Pennsylvania State University. Fellow in the American Society of Plant Biologists and the American Association for the Advancement of Science (AAAS).
Event: Robert D. Havener Seminar on “Innovations for Crop Productivity”.
http://ciat.cgiar.org/event/robert-d-havener-seminar-on-innovations-for-crop-productivity/
Access to large-scale omics datasets i.e. genomics, transcriptomics, proteomics, metabolomics, phenomics, etc. has revolutionized biology and led to the emergence of systems approaches to advance our understanding of biological processes. With decreasing time and cost to generate these datasets, omics data integration has created both exciting opportunities and immense challenges for biologists, computational biologists, biostatisticians and biomathematicians. Genomics, transcriptomics, proteomics, and metabolomics together they help to bring out the best of characters in plants.
Association mapping, also known as "linkage disequilibrium mapping", is a method of mapping quantitative trait loci (QTLs) that takes advantage of linkage disequilibrium to link phenotypes to genotypes.Varioius strategey involved in association mapping is discussed in this presentation
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.
Targeted Induced Local Lesions IN Genome. Mutations (Single base pair substitution) are created by traditionally used chemical mutagens. Identify SNPs and / or INDELS in a gene / genes of interest from a mutagenized population.
Majority of agronomic traits are quantitative and are controlled polygenetically.Instead of producing transgenic plants through single gene transfer many researchers are attempting on multigene engineering. The simultaneous transfer of multiple genes in to plants will enable us to produce plants with more desirable characters. Engineering of genes coding for complete metabolic pathways, bacterial operons or biopharmaceuticals that require an assembly of complex multisubunit proteins etc are some of the successful examples of multigene engineering.
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)
Current Status of TILLING and EcoTILLING:
TILLING and EcoTILLING technique have been adapted in diverse species including rice, maize, Lotus, poplar, Arabidopsis, wheat, barley, potato, tomato, sunflower, common bean, Field Mustard, clover, melon, pea, peanut, sorghum, rapeseed, soybean, melon, poplar, sugarcane, brassica and other for the purpose of gene detection, functional genomics, polymorphism assessment, plant breeding as described in case study part.
Ecotilling:
EcoTILLING is similar to TILLING, except that its objective is to identify natural genetic variation as opposed to induced mutations.
Many species are not amenable to chemical mutagenesis; therefore, EcoTILLING can aid in the discovery of natural variants and their putative gene function
This approach allows one to rapidly screen through many samples with a gene of interest to identify naturally occurring SNPs and / or small INs/DELS
iTILLING:
A new approach to the TILLING method that reduces costs and the time necessary to carry out mutation screening was developed for Arabidopsis and it is called iTILLING, individualized TILLING
A powerful non-transgenic reverse genetics method that combines chemical mutagenesis with PCR based screening to identify point mutations in regions of interest.
EcoTILLING is a molecular technique that is similar to TILLING, except that its objective is to uncover natural genetic variation as opposed to induced mutations.
As a periodontist, it is of utmost importance to understand the genetic basis of inheritance in periodontal diseases be able to relate to the various polymorphisms associated with periodontal diseases. This ppt presents the basics of genetics from the point of view of future understanding of polymorphisms related to periodontal diseases.
Gene trapping is a type of insertional mutagenesis that disrupts gene function by the integration of a vector in the intergeneric sequences. It provides an important and unique method for studying the relationship between gene expression and function and to characterize novel genes and analyze their importance in biological phenomena. It is insertional based gene discovery that utilizes random integration of reporter gene construct into genome and produce dominant expression phenotype.
It is performed with gene trap vectors that simultaneously mutate and report the expression of the endogenous gene at the site of insertion. The transformed cells are selected on the basis of selectable markers. Insertion events are detected and the trapped lines are established. This technique has been used to identify tissue specific and temporally regulated genes in plants and mice. It proves to be a powerful tool of functional genomics.
Size is important for cell function and is characteristic of different species and of different cell types in the same organism. How cell size is specified has intrigued researchers ever since cells became visible to biologists. Cell size determines the geometry of all intracellular compartments and sets the scale of biosynthetic processes. Biosynthesis increases in larger cells, which have proportionally more protein, total RNA, total mRNA, and mRNA for specific investigated genes.Plants are a good multicellular system for studying the regulation of cell size. As plant cells cannot migrate because they are encased in a cell wall matrix, their growth and division can be readily tracked and measured by live cell imaging and three-dimensional (3D) image analysis. Plants are also particularly relevant for studying how cell size relates to ploidy and to organ size because of the prominent role of polyploidy in plant evolution, plant development, and crop domestication.
ROLE OF CYTOKININS IN RETARDING LEAF SENESCENCEsukruthaa
Cytokinins are involved in the control of numerous and important processes associated with plant growth and development. They take part in the control of cell division, chloroplast development, bud differentiation, shoot initiation and growth or leaf senescence.
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.
Earliest Galaxies in the JADES Origins Field: Luminosity Function and Cosmic ...Sérgio Sacani
We characterize the earliest galaxy population in the JADES Origins Field (JOF), the deepest
imaging field observed with JWST. We make use of the ancillary Hubble optical images (5 filters
spanning 0.4−0.9µm) and novel JWST images with 14 filters spanning 0.8−5µm, including 7 mediumband filters, and reaching total exposure times of up to 46 hours per filter. We combine all our data
at > 2.3µm to construct an ultradeep image, reaching as deep as ≈ 31.4 AB mag in the stack and
30.3-31.0 AB mag (5σ, r = 0.1” circular aperture) in individual filters. We measure photometric
redshifts and use robust selection criteria to identify a sample of eight galaxy candidates at redshifts
z = 11.5 − 15. These objects show compact half-light radii of R1/2 ∼ 50 − 200pc, stellar masses of
M⋆ ∼ 107−108M⊙, and star-formation rates of SFR ∼ 0.1−1 M⊙ yr−1
. Our search finds no candidates
at 15 < z < 20, placing upper limits at these redshifts. We develop a forward modeling approach to
infer the properties of the evolving luminosity function without binning in redshift or luminosity that
marginalizes over the photometric redshift uncertainty of our candidate galaxies and incorporates the
impact of non-detections. We find a z = 12 luminosity function in good agreement with prior results,
and that the luminosity function normalization and UV luminosity density decline by a factor of ∼ 2.5
from z = 12 to z = 14. We discuss the possible implications of our results in the context of theoretical
models for evolution of the dark matter halo mass function.
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 .
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
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.
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.
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.
2. Intro..
• Epigenetics means ‘above’ or ‘on top of genetics’
Def :
The term epigenetics refers to heritable changes in gene expression that
does not involve changes to the underlying DNA sequence; a change in
phenotype without a change in genotype.
3. ⚫Conrad Waddington (1942) coined
the term, “epigenetic”. He is known
as father of epigenetics.
⚫Waddington’s attempted to explain
how a static set of DNA sequences
could dynamically give rise to a
complex organism.
4. Epigenetics describes phenomenon in which genetically identical
cells or organisms express their genomes differently, causing
phenotypic differences
Genetically identical
cells or individuals
Different epigenetic
modifications leading to
different expression
patterns
Different
phenotypes
5. Science7 April 2000:V
ol. 288. no. 5463, p. 38
Was Lamarck Just a Little Bit Right?
Michael Balter
Although Jean-Baptiste Lamarck is remembered mostly for the discredited theory
that acquired traits can be passed down to offspring, new findings in the field of
epigenetics, the study of changes in genetic expression that are not linked to
alterations in DNA sequences, are returning his name to the scientific literature.
Although these new findings do not support Lamarck's overall concept, they raise
the possibility that "epimutations," as they are called, could play a role in
evolution.
Lamarck was a true pioneer of evolutionary theory!
7
6. So the Epigenetics……
The fact that non-genetic variations that are obtained during an organism’s
life can be possibly be passed on to that organism’s offspring.
7. Epigenome
o An epigenome consists of a record of the chemical changes to the DNA
and histone proteins of an organism.
o These changes can be passed down to an organism's offspring.
o Changes in the epigenome can result in changes to the structure of
chromatin and changes to the function of the genome.
o The epigenome is a multitude of chemical compounds that can tell the
genome what to do.
Epialleles
• Alleles of a locus which have identical DNA sequences but display
different epigenetic states and which have been proposed to influence a
variety of phenotypes in plants and animals.
8. Genetics Epigenetics
mutations
alterations
Genetics vs Epigenetics
Changes in gene expression and/or function
and new phenotypes
Study of heredity and
variation of inherited
characters
Studies the structure
,interactions,function and
alterations of genes of
particular organism
Combination of alleles in
particular organism is
studied
Study of inheritable
changes caused by
modification of gene
expression
Different patterns of
methylation and
acetylation of DNA
and chromatin are
studied
www.pedia.com
10. Molecular epigenetic mechanism
Development of multicellular organisms occurs due to cells differentiation by various
programs of gene expression
Cells have own epigenetic signatures like
1. Genotype
2. Developmental history
3. Environmental influences and it is ultimately reflected in the phenotype of the cells and
the organism
DNA
Methylation
Histone
Modification
RNA
Interference
Gene
expression
12. 1.DNA Methylation
• A conserved epigenetic modification resulting from
the enzymatic addition of a methyl moiety to DNA
(i.e.. to Cytosine)
• Preferentially targeted to repetitive DNA associated
with heterochromatin
• Most common at symmetric cytosines (CpG,
CpNpG) in plants
• CpG sites or CG sites are regions of DNA where
a cytosine nucleotide is followed by
a guanine nucleotide in the
linear sequence of bases along its 5' → 3'
direction.
• CpG sites occur with high frequency in genomic
regions called CpG islands
13. Classes of methyl transferase
Enzymes that create new methylation
mark on DNA
Recognizes the methylation marks on the
parental strand of DNA and transfers new
methylation to the daughter strands after
DNA replication
**DRM - Domain Rearranged Methylase
MET -Methyl Transferase
CMT- Cytosine 5 Methyl transferase
14. Determination of DNA methylation
High-performance capillary Electrophoresis (HPCE)
Bisulfite treatment method
Methylation-sensitive representational difference analysis (MS-RDA)
15. High-performance
capillary Electrophoresis
(HPCE)
Cytosine and 5-methylcytosine can be identified
and quantified
Products are then separated by standard reverse
phase HPLC
Deoxyribonucleotides are further converted into
deoxyribonucleosides by treatment with alkaline
phosphatase
Total genomic DNA is hydrolyzed to
deoxyribonucleotides using a combination of
deoxyribonuclease and nuclease P1.
To measure global methylation level
through the quantification of 2’-
deoxynucleosides
Detection and quantification of 5-
methyl 2’-deoxycytidine in genomic
DNA is performed using micellar
HPCE with (UV/VIS)
Spectrophotometry detection
16. Bisulfite treatment
method
The PCR product is sequenced
DNA treated with bisulfite & all cytosine residues in
the DNA will be converted to uracil, the majority of 5-
methylcytosines remained intact
Restriction digestion products are run and DNA of
size range of interest is purified
DNA is digested into linear fragments that are more
readily denatured for bisulfite conversion than total
genomic DNA
Unmethylated cytosine is converted
to uracil while methylcytosine
remains unaltered.
In a polymerase chain reaction
(PCR),uracil is a template like
thymidine and all original cytosine
residues are converted to
thymidine.
PCR products are sequenced and
mapped onto genome data in order
to locate unchanged cytosine,which
is the methylcytosine.
18. Methylation-sensitive representational difference
analysis (MS-RDA)
Representational Difference Analysis (RDA) is a technique to find sequence
differences in two genomic or cDNA samples
Genomes or cDNA sequences from two samples are PCR amplified and differences
analyzed using subtractive DNA hybridization
This technique needs some methylation-sensitive enzymes such as HpaII and Msp1.
Genomic DNA is first digested with a methylation-sensitive restriction enzyme such
as HpaII and then a methylation insensitive isoschizomer MspI is used in a parallel
digestion reaction.
Subsequently, these fragments are selectively amplified by fluorescently labelled
primers.
PCR products from different individuals are compared and once an polymorphic locus
is recognized, the desired DNA fragment can be isolated from a denaturing
polyacrylamide gel and sequenced to check methylation pattern.
19. Histone Modification
• They are the chief protein components of chromatin, acting as spools
around which DNA winds, and playing a role in gene regulation.
⚫Methylation
⚫Acetylation
⚫Phosphorylation
Combinations of Histone
modifications constitute
‘Histone Code
20. A. Histone methylation
It is a process by which methyl
groups are transferred to amino
acids of histone proteins that make
up nucleosomes
Enzyme required
Histone methytransferases (HMTs)
LMT- Lysine methyl transferase
Methylation can result in activation or
repression of genes.
21. B. Histone Acetylation & Deacetylation
Histone acetylation
– Histone acetyl transferases (HATs)
Adds acetyl groups to histone tails.
Reduces positive charge and weakens interaction of histones with DNA
Facilitates transcription by making DNA more accessible to RNApolymerase II
Histone deacetylation
– Histone deacetylases (HDACs)
Removes acetyl groups from histone tails
Increases interaction of DNAand histones
Represses transcription
22. C. Histone Phosphorylation
Phosphorylation
Enzyme required – Protein kinase
Phosphorylation increase the negative charge on Histone as a result less interaction
between DNA and histones that leads to chromatin de-condensation.
Dephosphorylation
Enzyme required - phosphatase
increase positive charge followed by chromatin condensation
23. 3. RNAi (RNA INTERFERENCE)
• Also called post transcriptional gene silencing (PTGS)
• Is a RNA- dependent gene silencing process
in which RNA molecules inhibit gene action
• Andrew Fire and Craig C. Mello shared the Nobel Prize in
Physiology and Medicine – 2006
• RNAi involves small RNA molecules called short interfering
RNAs (si RNA) and micro RNAs (mi RNA).
• These molecules are 21 to 28 base pairs long and produced
from larger dsRNA molecules by enzyme called dicer.
24. Function of both species is regulation of gene expression.
Difference is in where they originated. siRNA originated from dsRNA
and miRNA originated with ssRNA that form hairpin secondary
structure.
siRNA is most commonly a response to foreign RNA (usually viral) and
is often 100% complementary to the target.
miRNA regulate post transcriptional gene expression and often not 100%
complementary to the target.
26. Detection
• Isolate total RNA, separate small RNA fraction on PAGE, reverse transcribe, clone
and sequence – biochemical approach
• Identify miRNAs based on complementarity with targeted sequences by
computational methods– in silico approach
• The microRNA Registry v2.0 contains 506 miRNA entries from six organisms
including rice, maize, soyabean, sugarcane, sorghum etc.
(http://www.sanger.ac.uk/Software/Rfam/mirna/) (Griffith-Jones, 2004, Nucleic Acids
Res. 32D:109-111)
• MIRO – computational tool to identify miRNAs and their targets in plants
28. A. Epigenetic variation
• Plant populations show phenotypic diversity, which may be caused by genetic
and epigenetic variation.
• It has recently been shown that new epigenetic variants are generated at a higher
rate than genetic variants.
• Epimutant - If an epigenetic variant or epiallele has a phenotypic effect and is
more or less stably inherited to the progeny- create new sources of variation
• Dedifferentiation and redifferentiation in tissue culture results in modification of
epigenetic patterns leading to Somaclonal variations
• Rice mutant line Low Glutelin Content-1 – First commercial cultivar showing RNAi
This dominant mutation in glutelin gene produces a hp RNA and via RNAi
reduces glutelin content
29. B. Plant stress Tolerance
Abiotic
stress
Crop Response References
Salt Wheat Lower methylation level in
tolerant cultivar
Wang et al., 2014
Heat Grapevine Transgenerational inheritance of
methylation after removal of stress
Baranek et al., 2015
Cold Maize Increase in H3K9ac and H4K5ac in
promoter of cell cycle genes
Zhao et al., 2014
Heavy metal Chickpea Hypomethylation in tolerant
upon prolonged exposure
Rakei et al., 2015
30. Biotic stress
Example: Georg Jander (2007) exposed Arabidopsis plants to caterpillars and
tested what effect this had on the plants’offspring.
The second generation, he found that If attacked by caterpillars, offspring's
responded more strongly—reducing caterpillar growth by 40% compared to
caterpillars feeding on control plants.
31. C. Yield and Heterosis
• Heterosis involves change in global gene expression (Tsaftaris, 1995)
• Non additive gene expression observed in hybrids due to regulatory
interactions
• Zein expression in Maize endosperms (Song and Messing, 2003)
• Protein levels in Maize root tips (Romagnoli, 1990)
• Significant differences found in parental alleles in maize hybrids to
that in parents due to differences in transcriptional regulation (Guo et al.,
2004)
• Hybrids are more resistant to site specific methylation changes under
stress leading to increased stability (Kovacevic et al., 2005)
32. D. Parental Imprinting
• Process by which only one of the parental alleles either maternal or
paternal is active in an offspring
• Mitotically stable epigenetic modification inactivates one of the parent
alleles
• MEA/FIE (FERTILIZATION-INDEPENDENT ENDOSPERM) /FIS2 (FERTILISATION-INDEPENDENT SEED)
PcG (Polycomb group) like complex plays an important role in seed
development and shows imprinting (Baroux et al., 2002, Adv.Genet.
46:165-214)
• Imprinting is by maternal specific removal of methylation by DME DNA
glycosylase in female gametophyte
33. E. Plant Resistance to Viruses and other Pathogens
• PTGS based viral resistance does not involve transgenic production of
viral genes or proteins, nor transgenic RNA (Goldbach et al. 2003)
• Viral Induced Gene silencing VIGS used for characterisation of genes
associated with local and systemic resistance in barley and other
cereals
34. F. Rnai IN PLANT BREEDING
• More efficient than antisense based silencing
• Silencing achieved by using a transgene producing hairpin RNA
with dsRNA region
• Particularly useful in silencing genes in polyploids or genes
belonging to multigene families
• Zein-a protein repression achieved in maize RNAi to increase
lysine content without the complications associated with opaque-
2 mutant (Segal et al., 2003)
• Caffeine synthesising enzyme expression repressed by RNAi to
reduce caffeine content by 50-70% (Ogita et al., 2003)
• RNAi silencing suppression seen in viruses can be used for
overproduction of desired proteins in plants
35.
36. The field of epigenetics has rapidly developed into one of the most influential areas of
scientific research.
Recent advances in analytical methodology have allowed for a significant expansion of
what is known about genome wide mapping of DNA methylation and histone
modifications.
Good knowledge of epigenetic mechanisms leads to better understanding of
regulation of gene expression at transcriptional and post-transcriptional levels.
Epigenetic mechanisms such as DNA methylation and histone modification play a key
role in plant development and stress response.
Epigenetics is involved in a number of biological phenomena including developmental
control in plants
Conclusion
37. References
• Balter,M. 2000. GENETICS: Was Lamarck Just a Little Bit Right? Science, 288(5463), 38. doi:10.1126/science.288.5463.38
• Difference between genetics and epigenetics - www.pedia.com
• Tsaftaris et al., 2005. Epigenetic mechanisms in plants and their implications in plant breeding.
• Wang et al., 2014. Induced and Constitutive DNA Methylation in a Salinity-Tolerant Wheat Introgression Line. Plant Cell
Physiology. 55(7): 1354–1365. doi:10.1093/pcp/pcu059
• Baroux et al., 2002. Genomic Imprinting During Seed Development. Advances in Genetics. 46:165-214.
https://doi.org/10.1016/S0065-2660(02)46007-5
