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.
i explained about basics of genome engineering and crispr system.
CRISPR will change the world and it is just the beginning, are you ready to meet the future? you think its great and beautiful or.....?
please give your feedback to my email
pooyanaghshbandi@yahoo.com
i am starting to write a critical and fantastic review article about CRISPR, if you are interested to join please contact me.
This presentation highlights the basics and application of genome editing strategies in plants, strategies to reduce off-target mutation, identification of mutant analysis etc.
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.
i explained about basics of genome engineering and crispr system.
CRISPR will change the world and it is just the beginning, are you ready to meet the future? you think its great and beautiful or.....?
please give your feedback to my email
pooyanaghshbandi@yahoo.com
i am starting to write a critical and fantastic review article about CRISPR, if you are interested to join please contact me.
This presentation highlights the basics and application of genome editing strategies in plants, strategies to reduce off-target mutation, identification of mutant analysis etc.
Genome editing with the CRISPR-Cas9 system has become one of the major tools in modern biotechnology. This slide share discusses the fundamentals in a simple, easy to understand format.
Genome Editing Techniques by Kainat RamzanKainatRamzan3
Genome technology has revolutionized biological science through techniques of Gene Editing in order to edit any organism's genome.MegNs and zinc-finger nucleases are commonly understood to be used, as is the effector's transcriptional activator-like nucleases. In CRISPR/Cas9, genetic alterations, and gene functionality have become a well-known tool for understanding gene targeting.
The next generation of crispr–cas technologies and Applicationsiqraakbar8
The prokaryote-derived CRISPR–Cas genome editing systems have transformed our ability to manipulate, detect, image and annotate specific DNA and RNA sequences in living cells of diverse species. The ease of use and robustness of this technology have revolutionized genome editing for research ranging from fundamental science to translational medicine. Initial successes have inspired efforts to discover new systems for targeting and manipulating nucleic acids, including those from Cas9, Cas12, Cascade and Cas13 orthologues.
A CRISPR/Cas9, works like a biological version of a word-processing programme’s “find and replace”. Its simplicity and extremely low cost of implementation is the reason to use. How Cas 9 is activated and its mechanism (DNA binding and cleavage), it's regulation and application in human disease therapy, new drug screening, agriculture and biofuel etc.
Now a day's these technique is tremendously use for in lab by using foreign Dna to to producing insulin in bacteria , plant with high yielding capacity by using Gene from another species
The Evolution of Science Education PraxiLabs’ Vision- Presentation (2).pdfmediapraxi
The rise of virtual labs has been a key tool in universities and schools, enhancing active learning and student engagement.
💥 Let’s dive into the future of science and shed light on PraxiLabs’ crucial role in transforming this field!
Genome editing with the CRISPR-Cas9 system has become one of the major tools in modern biotechnology. This slide share discusses the fundamentals in a simple, easy to understand format.
Genome Editing Techniques by Kainat RamzanKainatRamzan3
Genome technology has revolutionized biological science through techniques of Gene Editing in order to edit any organism's genome.MegNs and zinc-finger nucleases are commonly understood to be used, as is the effector's transcriptional activator-like nucleases. In CRISPR/Cas9, genetic alterations, and gene functionality have become a well-known tool for understanding gene targeting.
The next generation of crispr–cas technologies and Applicationsiqraakbar8
The prokaryote-derived CRISPR–Cas genome editing systems have transformed our ability to manipulate, detect, image and annotate specific DNA and RNA sequences in living cells of diverse species. The ease of use and robustness of this technology have revolutionized genome editing for research ranging from fundamental science to translational medicine. Initial successes have inspired efforts to discover new systems for targeting and manipulating nucleic acids, including those from Cas9, Cas12, Cascade and Cas13 orthologues.
A CRISPR/Cas9, works like a biological version of a word-processing programme’s “find and replace”. Its simplicity and extremely low cost of implementation is the reason to use. How Cas 9 is activated and its mechanism (DNA binding and cleavage), it's regulation and application in human disease therapy, new drug screening, agriculture and biofuel etc.
Now a day's these technique is tremendously use for in lab by using foreign Dna to to producing insulin in bacteria , plant with high yielding capacity by using Gene from another species
The Evolution of Science Education PraxiLabs’ Vision- Presentation (2).pdfmediapraxi
The rise of virtual labs has been a key tool in universities and schools, enhancing active learning and student engagement.
💥 Let’s dive into the future of science and shed light on PraxiLabs’ crucial role in transforming this field!
This presentation explores a brief idea about the structural and functional attributes of nucleotides, the structure and function of genetic materials along with the impact of UV rays and pH upon them.
ESR spectroscopy in liquid food and beverages.pptxPRIYANKA PATEL
With increasing population, people need to rely on packaged food stuffs. Packaging of food materials requires the preservation of food. There are various methods for the treatment of food to preserve them and irradiation treatment of food is one of them. It is the most common and the most harmless method for the food preservation as it does not alter the necessary micronutrients of food materials. Although irradiated food doesn’t cause any harm to the human health but still the quality assessment of food is required to provide consumers with necessary information about the food. ESR spectroscopy is the most sophisticated way to investigate the quality of the food and the free radicals induced during the processing of the food. ESR spin trapping technique is useful for the detection of highly unstable radicals in the food. The antioxidant capability of liquid food and beverages in mainly performed by spin trapping technique.
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.
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/
ANAMOLOUS SECONDARY GROWTH IN DICOT ROOTS.pptxRASHMI M G
Abnormal or anomalous secondary growth in plants. It defines secondary growth as an increase in plant girth due to vascular cambium or cork cambium. Anomalous secondary growth does not follow the normal pattern of a single vascular cambium producing xylem internally and phloem externally.
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.
Toxic effects of heavy metals : Lead and Arsenicsanjana502982
Heavy metals are naturally occuring metallic chemical elements that have relatively high density, and are toxic at even low concentrations. All toxic metals are termed as heavy metals irrespective of their atomic mass and density, eg. arsenic, lead, mercury, cadmium, thallium, chromium, etc.
Remote Sensing and Computational, Evolutionary, Supercomputing, and Intellige...University of Maribor
Slides from talk:
Aleš Zamuda: Remote Sensing and Computational, Evolutionary, Supercomputing, and Intelligent Systems.
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Inter-Society Networking Panel GRSS/MTT-S/CIS Panel Session: Promoting Connection and Cooperation
https://www.etran.rs/2024/en/home-english/
2. What is gene editing?
• Making precise alterations to the genome
• Removal of DNA
• Insertion of DNA
• Base pair alterations of the DNA
3. How do we edit the genome?
• At the heart of genome editing is the ability to make a precise cut at a
specific sequence within the genome
Creation of frameshifts
Insertion of DNA
Alterations to the DNA sequence
Removal of DNA
Requires the activity of an endonuclease
4. HDR for genome editing
An Acad Bras Cienc. 2015 Aug;87(2 Suppl):1323-48.
Initial gene targeting utilized homologous recombination to
insert DNA into a specific locus.
This process is extremely inefficient.
Reprod Biol Endocrinol. 2014 Nov 24;12:108.
The process can be made more efficient by
cutting the DNA and stimulating homology
directed repair
5. Zinc Finger Nucleases: Targeting the Genome
The first specific mechanism to drive a nuclease to a
specific region of the genome.
Made by fusing the Fok I endonuclease domain with
DNA binding zinc fingers.
Zinc fingers are designed in modules of 2, each
recognizing 6 bps. These modules can be strung
together to generate a zinc finger protein that will
recognize as specific sequence in the DNA ( ~24 bp).
Fok 1 is a non-specific endonuclease that has to
dimerize in order to cleave the DNA.
You need to 2 Zinc Finger/Fok1 molecules, each
targeting opposite strands to create a very specific
cleavage site in the DNA.
7. Zinc Finger Nucleases are not very practical
Designing Zinc Finger nucleases with proper specificity is challenging and requires expertise.
Furthermore, zinc finger nucleases have been patented limiting the competition of commercial providers.
Cost of getting a set of zinc finger nucleases is $3000-$7000.
8. TALENs
• Transcription activator-like effector nucleases
J Clin Invest. 2014;124(10):4154-4161
Uses the same conceptual approach as zinc
finger nucleases but replaced the DNA
binding activity of zinc fingers with tale
proteins
9. TALE proteins have a code of specificity
Nature Biotechnology 29, 143–148 (2011)
TALE proteins are bacterial transcriptional activators used to
regulate genes in plants the bacteria infect.
These TALE proteins have a repeat domain of 33-35 bp. Each
repeat recognizes a single base.
A dinucleotide sequence in the middle of the repeat dictates
the recognized base.
10. TALENs increased the practical usage of gene
targeting
Not patented.
Labs generated kits of the different targeting
repeats that could be easily assembled.
No specialized expertise needed for design.
Could be assembled in any lab at relatively low
cost.
Still need to make two unique protein
encoding plasmids for each site you want to
target.
11. CRISPR revolution- hijacking the bacterial
immune system
Foreign DNA will get inserted into a specific locus of the bacterial
genome called the Clustered Regularly Interspaced Short
Palindromic Repeats (CRISPR). This locus represents a series of short
repeats broken up by spacers. The spacers are actually foreign
pieces of DNA that have previously integrated into the locus.
The CRISPR locus produces a transcript which gets processed to
create small RNA molecules that contain the spacer and a repeat.
This RNA is called the crRNA.
For type II CRISPR systems, the crRNA interacts with a second RNA
called the tracrRNA and the Cas9 protein.
Together this complex uses the spacer RNA sequence to guide the
complex to foreign DNA. The Cas9 protein then cleaves the DNA
Biochimie.Volume 117, October 2015, Pages 119-128
12. CRISPR targets a DNA sequence
Through RNA/DNA base pairing- the spacer element
interacts with DNA with complimentary sequence
forming an R-loop. An R-loop is three standed
nucleic acid with an RNA-DNA compliment and a
single stranded DNA.
The tracrRNA is needed to form a complex with Cas9
Cas9 is an endonuclease which has two separate
endonuclease domains (HNH & RuvC) which cleave
each strand of the DNA separately.
The carboxy terminus of Cas9 must interact with a
specific DNA element to become activated called the
PAM site.
The PAM site consists of an NGG.
Volume 23, Issue 4, p225–232, April 2015
13. Utilizing this system to target DNA cleavage
There are 3 components to bacterial Cas9/CRISPR system:
1) crRNA- the only part that changes between target sites
2) tracrRNA
3) Cas9
The Spacer sequence is the only part that needs to be
adjusted to target Cas9 cleavage
14. Simplifying the systems into two modules
Combining the crRNA and tracrRNA into one RNA called the
single guide RNA (sgRNA).
Simply turned the junction point into a hairpin.
It can now be encoded by one gene
Molecular Cell. Volume 56, Issue 2, p333–339, 23 October 2014
15. The CRISPR/Cas9 system makes it incredibly
easy to target a nuclease to DNA
Several systems developed in which small
oligos can be easily cloned into a plasmid to
make a guide RNA.
Some of these systems already have Cas9
expression cassettes making it a 1 plasmid
system.
Easy 1 step cloning.
Inexpensive and quick
Target anywhere in the genome with an NGG
site.
All things being random, should be present
every 16 bp
In humans- averages to every 42 bp.
https://www.biocat.com/genomics/genome-engineering/crispr-cas9-smartnuclease-genome-engineering-system-
vector-based
16. The spacer region of sgRNA does not need
100 % base pairing
Nature Methods 12, 1150–1156 (2015)
Similar to miRNA/siRNA, there is a seed sequence that is
more critical in driving the specificity of the complex to
targets
17. Different Cas proteins can modify gene
targeting
Increases potential cut sites (ex: Cas9 needs
GG, Cas12a needs YTT)
Potentially better recombination efficiency
with Cas12a as the cut site is not in the
seed region
Can target RNA.
18. CRISPR has off target
effects
Nature Biotechnology 33, 187–197 (2015)
CRISPR recruits Cas9 to potentially hundreds of sites
across the genome
Read counts suggesting how often
these off target sites are hit
19. Using a nickase to drive HDR with less off
target effects
Cell. Volume 154, Issue 6, p1380–1389, 12 September 2013
Mutations (D10A) to HNH function of Cas9 cause single strand nicks in the
DNA.
Using two sgRNAs to drive Cas9D10A to two close sites can cause a double
strand break.
20. CRISPR initiates indels at target sites
https://www.addgene.org/crispr/guide/
Cas9 mediated DNA cleavage is often repaired by NHEJ.
Using the CRISPR/Cas9 systems, this almost always results an
insertion or deletion of base pairs at the cut site.
When targeted to coding regions, often results in frameshifts that can
cause knockout of the targeted gene if done in the proper region of
the cds.
21. NHEJ competes with HDR to limit the
efficiency of gene editing
Biology Open 2014 3: 271-280;
The inefficiency of HDR in gene editing serves as a major
road block for its use in therapy and animal generation
22. Improving CRISPR through fusion proteins
Fusion of Cas9 to hRad51 mutants improves homology directed repair.
hRad51 directs HDR to single strand DNA (improves HDR at nicked sites).
Mutants can limit “perfect repair”
23. CRISPR Prime to improve editing
CRISPR Prime uses the Cas9 nickase fused to a reverse transcriptase
domain along with a special guide RNA called a pegRNA. The pegRNA is
a guide domain along with a template to modify the cut site.
The 5’ Flap gets chewed back by enzymes associated with DNA
replication (okazaki fragments).
Improved efficiency with lower off target effects. Efficiencies as high as
40%.
Nature. 2019 Oct 21.
Use an additional gRNA to nick non-edited
strand.
24. Using CRISPR to do more than edit the
genome
dCas9 is a version of the Cas9 that has no
endonuclease activity
dCas9 can serve has a protein cargo ship for fusion
partners to modify epigenome or act as a steric
hindrance to DNA binding.
Integr Biol (Camb). 2017 Feb 20;9(2):109-122.
25. dCas9 to regulate transcription
Fusions of dCas9 to KRAB transcriptional repressors
or VP16 transcriptional activators allows for precise
targeting to almost anywhere in the genome.
26. dCas9 can be used to edit the epigenome
Fusions of dCas9 to either
tet or DNMT enzymes
allows for epigenomic
editing
Cell. Volume 167, Issue 1, p233–247.e17, 22 September 2016
27. dCas9 DNMT3 can target methylation
Cell. Volume 167, Issue 1, p233–247.e17, 22 September 2016
The locus only the presence of Dox (inducible system) and
the proper sgRNA is able to drive methylation.
28. Base Editing- CRISPR fusion to base modifying
enzymes
C→T, G→A, A→G, and T→C
Four type of base editing tools
Uses deamination to convert C-T using
APOBEC protein.
Or
Uses TadA to convert A to I(G)
Science 27 Oct 2017:
Vol. 358, Issue 6362, pp. 432-433
29. Directing base editing using
different fusion proteins
Different enzyme variants have different target preferences within the R-
loop.
Use of the PAM site and the enzyme directs which base pairs are most likely
to be edited