Here recent study by Dienstbier performs a comparison across next generation sequencing using RNA Seq analysis and DNA microarray to establish regulatory action for Hfq protein in B. pertussis.
Codon optimization is one of the key steps in achieving the high level expression of target gene. There are some key factors for consideration including transcription efficiency, translation efficiency, gene synthesis and protein folding.
Codon optimization is one of the key steps in achieving the high level expression of target gene. There are some key factors for consideration including transcription efficiency, translation efficiency, gene synthesis and protein folding.
pBluescript is an example of a combination between plasmids and phages (phagemids).
Phagemids represent a hybrid type of class of vectors that serve to produce single-stranded DNA.
Replication,transcription,translation complete the central dogma of life.How mRNA,tRNA,rRNA act on ribosomes for protein synthesis.Difference between eukaryotes and prokaryotes
REGULATION OF
GENE EXPRESSION
IN PROKARYOTES & EUKARYOTES .
This presentation is enriched with lots of information of gene expression with many pictures so that anyone can understand gene expression easily.
Gene expression is the process by which the information encoded in a gene is used to direct the assembly of a protein molecule.
Gene expression is explored through a study of protein structure and function, transcription and translation, differentiation and stem cells.
It is the process by which information from a gene is used in the synthesis of a functional gene product.
These products are often proteins, but in non-protein coding genes such as ribosomal RNA (rRNA), transfer RNA (tRNA) or small nuclear RNA (snRNA) genes, the product is a functional RNA.
The process of gene expression is used by all known life - eukaryotes (including multicellular organisms), prokaryotes (bacteria and archaea)
Regulation of gene expression:
Regulation of gene expression includes a wide range of mechanisms that are used by cells to increase or decrease the production of specific gene products (protein or RNA).
Gene regulation is essential for viruses, prokaryotes and eukaryotes as it increases the versatility and adaptability of an organism by allowing the cell to express protein when needed.
CLASSIFICATION OF GENE WITH RESPECT TO THEIR EXPRESSION:
Constitutive ( house keeping) genes:
Are expressed at a fixed rate, irrespective to the cell condition.
Their structure is simpler.
Controllable genes:
Are expressed only as needed. Their amount may increase or decrease with respect to their basal level in different condition.
Their structure is relatively complicated with some response elements.
TYPES OF REGULATION OF GENE:
positive & negative regulation.
Steps involving gene regulation of prokaryotes & eukaryotes.
Operon-structure,classification of mechanisms- lac operon,tryptophan operon ,
and many things related to gene expression.
This is a video slide so anyone can understand this topic easily by seeing pictures included in this slide.
Manipulation of gene expression in prokaryotesSabahat Ali
For expression of gene in a particular vector, always used strong regulatable promoter (lac promoter, trp promoter, tac promoter , trc promoter, pL promoter, T7 gene promoter)
use of dual plasmid system & fusion proteins
How we can increase our protein product yield?
Gene rehulation in prokaryotes and eukaryotesSuresh Antre
Gene regulation drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms where the different types of cells may possess different gene expression profile.
2 a. RNA polymerase II that is actively transcribing a gene (activel.pdfabhinavbhatnagar201
2 a. RNA polymerase II that is actively transcribing a gene (actively progressing along the DNA
polymerizing an RNA molecule) is phosphorylated. Researchers evaluated the abundance (the
frequency of occupancy) of RNA PolII at difference positions along a gene, and correlated that
with the introns and exons of the gene. The figure represents a portion of one gene that is
transcribed from left to right. The X axis corresponds to position along the DNA, and the boxes
below the figure illustrate where the exons are (with the lines between them representing the
introns). The Y axis represents the relative frequency of occupancy (relative abundance of active
RNA PolII using two different measures [pSer2 and pSer5]). A high the value on this axis means
that, when they sample whether RNA polII is present at the given position, they find the enzyme
present more frequently than at positions where the value is low.
a. With respect to intron/exon boundaries, what positions does the (transcribing) RNA PolII
occupy most?
b. How do you interpret this observation (i.e., does it say anything about how fast or how slow
RNA PolII is moving at different positions across the gene)? Does it suggest anything about the
relationship between transcription and splicing?
2b. You characterize the sequence of a full-length cDNA and the corresponding genomic DNA
for a particular intron-containing gene from mouse cells. When you align them to each other
using a computer program, the exons of the cDNA align perfectly in some regions with pieces of
the genomic DNA, whereas other exons appear to have a small number of specific nucleotide
differences compared to the genomic DNA. Assume this genomic DNA and cDNA come from
the same individual, and there are no sequencing mistakes. What might account for these
differences? ,241 132 15 98 459605 719M850 1341/1417 Ratio pser relative to RNAPI
Solution
2a RNA polymerase II at first recognizes and binds to the promoter DNA forming a state termed
as the closed complex. Following this the DNA surrounding the transcription start site is
unwound and the template strand is positioned in the Polymerase active site, forming the open
complex. Transcription initiation then commences, initially producing short RNA products.
b Alternative splicing is a crucial mechanism for gene regulation and for generating genomic
diversity. Recent studies indicate that the expression of nearly 95% of human multi-exon genes
involves alternative splicing. In metazoans, alternative splicing plays an important role in
generating different protein products that function in diverse cellular processes including cell
growth, differentiation and death.
Splicing is carried out by the spliceosome. A massive structure in which five small nuclear
ribonucleoprotein particles (snRNPs) and a large number of auxiliary proteins cooperate to
accurately recognize the splice sites and catalyse the two steps of the splicing reaction.
2b There is consistent evidence that both proce.
pBluescript is an example of a combination between plasmids and phages (phagemids).
Phagemids represent a hybrid type of class of vectors that serve to produce single-stranded DNA.
Replication,transcription,translation complete the central dogma of life.How mRNA,tRNA,rRNA act on ribosomes for protein synthesis.Difference between eukaryotes and prokaryotes
REGULATION OF
GENE EXPRESSION
IN PROKARYOTES & EUKARYOTES .
This presentation is enriched with lots of information of gene expression with many pictures so that anyone can understand gene expression easily.
Gene expression is the process by which the information encoded in a gene is used to direct the assembly of a protein molecule.
Gene expression is explored through a study of protein structure and function, transcription and translation, differentiation and stem cells.
It is the process by which information from a gene is used in the synthesis of a functional gene product.
These products are often proteins, but in non-protein coding genes such as ribosomal RNA (rRNA), transfer RNA (tRNA) or small nuclear RNA (snRNA) genes, the product is a functional RNA.
The process of gene expression is used by all known life - eukaryotes (including multicellular organisms), prokaryotes (bacteria and archaea)
Regulation of gene expression:
Regulation of gene expression includes a wide range of mechanisms that are used by cells to increase or decrease the production of specific gene products (protein or RNA).
Gene regulation is essential for viruses, prokaryotes and eukaryotes as it increases the versatility and adaptability of an organism by allowing the cell to express protein when needed.
CLASSIFICATION OF GENE WITH RESPECT TO THEIR EXPRESSION:
Constitutive ( house keeping) genes:
Are expressed at a fixed rate, irrespective to the cell condition.
Their structure is simpler.
Controllable genes:
Are expressed only as needed. Their amount may increase or decrease with respect to their basal level in different condition.
Their structure is relatively complicated with some response elements.
TYPES OF REGULATION OF GENE:
positive & negative regulation.
Steps involving gene regulation of prokaryotes & eukaryotes.
Operon-structure,classification of mechanisms- lac operon,tryptophan operon ,
and many things related to gene expression.
This is a video slide so anyone can understand this topic easily by seeing pictures included in this slide.
Manipulation of gene expression in prokaryotesSabahat Ali
For expression of gene in a particular vector, always used strong regulatable promoter (lac promoter, trp promoter, tac promoter , trc promoter, pL promoter, T7 gene promoter)
use of dual plasmid system & fusion proteins
How we can increase our protein product yield?
Gene rehulation in prokaryotes and eukaryotesSuresh Antre
Gene regulation drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms where the different types of cells may possess different gene expression profile.
Similar to Differential Gene Expression Analysis of RNA Seq and Microarray Data highlights role of Hfq gene in expression regulation and parthenogenesis
2 a. RNA polymerase II that is actively transcribing a gene (activel.pdfabhinavbhatnagar201
2 a. RNA polymerase II that is actively transcribing a gene (actively progressing along the DNA
polymerizing an RNA molecule) is phosphorylated. Researchers evaluated the abundance (the
frequency of occupancy) of RNA PolII at difference positions along a gene, and correlated that
with the introns and exons of the gene. The figure represents a portion of one gene that is
transcribed from left to right. The X axis corresponds to position along the DNA, and the boxes
below the figure illustrate where the exons are (with the lines between them representing the
introns). The Y axis represents the relative frequency of occupancy (relative abundance of active
RNA PolII using two different measures [pSer2 and pSer5]). A high the value on this axis means
that, when they sample whether RNA polII is present at the given position, they find the enzyme
present more frequently than at positions where the value is low.
a. With respect to intron/exon boundaries, what positions does the (transcribing) RNA PolII
occupy most?
b. How do you interpret this observation (i.e., does it say anything about how fast or how slow
RNA PolII is moving at different positions across the gene)? Does it suggest anything about the
relationship between transcription and splicing?
2b. You characterize the sequence of a full-length cDNA and the corresponding genomic DNA
for a particular intron-containing gene from mouse cells. When you align them to each other
using a computer program, the exons of the cDNA align perfectly in some regions with pieces of
the genomic DNA, whereas other exons appear to have a small number of specific nucleotide
differences compared to the genomic DNA. Assume this genomic DNA and cDNA come from
the same individual, and there are no sequencing mistakes. What might account for these
differences? ,241 132 15 98 459605 719M850 1341/1417 Ratio pser relative to RNAPI
Solution
2a RNA polymerase II at first recognizes and binds to the promoter DNA forming a state termed
as the closed complex. Following this the DNA surrounding the transcription start site is
unwound and the template strand is positioned in the Polymerase active site, forming the open
complex. Transcription initiation then commences, initially producing short RNA products.
b Alternative splicing is a crucial mechanism for gene regulation and for generating genomic
diversity. Recent studies indicate that the expression of nearly 95% of human multi-exon genes
involves alternative splicing. In metazoans, alternative splicing plays an important role in
generating different protein products that function in diverse cellular processes including cell
growth, differentiation and death.
Splicing is carried out by the spliceosome. A massive structure in which five small nuclear
ribonucleoprotein particles (snRNPs) and a large number of auxiliary proteins cooperate to
accurately recognize the splice sites and catalyse the two steps of the splicing reaction.
2b There is consistent evidence that both proce.
1. Levels of gene regulation
The observation that differences in the RNA and protein content of different tissues are not paralleled by significant differences in their DNA content indicates that the process whereby DNA produces mRNA must be the level at which gene expression is regulated in eukaryotes. In bacteria this process involves only a single stage, that of transcription, in which RNA copy of the DNA is produced by the enzyme RNA polymerase. Even while this process is still occurring, ribosomes attach to the nascent RNA chain and begin to translate it into protein. Hence cases
of gene regulation in bacteria, such as the switching on of the synthesis of the enzyme β-galactosidase in response to the presence of lactose (its substrate), are mediated by increased transcription of the appropriate gene. Clearly, a similar regulation of gene transcription in different tissues, or in response to substances such as steroid hormones which induce the synthesis of new proteins, represents an attractive method of gene regulation in eukaryotes.
In contrast to the situation in bacteria, however, a number of stages intervene between the initial synthesis of the primary RNA transcript and the eventual production of mRNA (Fig. 1).
The initial transcript is modified at its 5′ end by the addition of a cap structure containing a modified guanosine residue and is subsequently cleaved near its 3′ end, followed by the addition of up to 200 adenosine residues in a process known as polyadenylation. Subsequently, intervening sequences or introns, which interrupt the protein-coding sequence in both the DNA and the primary transcript of many genes. Although this produces a functional mRNA, the spliced molecule must then be transported from the nucleus, where these processes occur, to the cytoplasm where it can be translated into protein.
1 At least 2 questions from this section will be on the .docxmercysuttle
1
At least 2 questions from this section will be on the final exam
SAMPLE QUESTIONS FOR THE FINAL EXAM
Question 1. Ferritin is a protein involved in the storage of iron inside cells. To prevent toxic accumulation of
too much iron inside cells, the intracellular level of ferritin is tightly regulated. To study the regulation of
ferritin synthesis, mammalian cells are grown with or without iron in the culture medium. Note that iron in the
culture medium is rapidly transported inside cells.
a) Upon addition of iron to the culture medium, the intracellular concentration of ferritin mRNA is unchanged
but the concentration of ferritin protein increases. How do you think ferritin expression is regulated? Briefly
explain.
The regulatory sequence given below is found in the ferritin mRNA between the cap structure and the start
codon.
5’-GGGUUUCCGUUCAACAGUGCUUGGACGGAAACCC-3’
Mutations within in this sequence are used to study the regulation of ferritin expression. The following
observation are made:
• ferritin expression is high, independent of the iron concentration, when (i) the entire region is deleted, or
(ii) the region located upstream of the underlined sequence is deleted or (iii) the underlined sequence is
replaced with a random sequence.
• ferritin expression remains iron-dependent when this region is replaced by the following sequence:
5’-GGGCUCAGGUUCAACAGUGCUUGGACCUGAGCCC-3’.
Note that the sequence differences are indicated in bold.
b) Explain why these observations suggest that both sequence and structure of the 5’ end of ferritin mRNA are
important for the regulation of ferritin expression.
c) Ferritin translation becomes iron-independent when the regulatory sequence is moved from the 5’ side
(upstream of the open reading frame) to the 3’ side (downstream of the open reading frame) of ferritin mRNA.
Which step of ferritin translation do you think is affected by the intracellular level of iron?
d) IRP is a protein involved in the regulation of ferritin expression. Anti-IRP antibodies attached to sepharose
beads are added to a cell extract, then the extract is centrifuged to separate the pellet fraction (containing the
sepharose beads ) from the supernatant fraction.
If the cells are cultured in the absence of iron, ferritin mRNA is found together with IRP in the pellet. In
contrast when cells are cultured in the presence of iron ferritin mRNA remains in the supernatant fraction while
IRP alone is found in the pellet. Briefly explain the likely role of IRP in the regulation of ferritin expression.
Question 2. You are studying the development of a newly discovered insect. Like drosophila, it undergoes a
stage in early larval development where the eve gene is expressed in a pattern of 7 stripes. You are particularly
interested in stripes 2 and 5. The following figures show the organization of the cis-acting elements that control
the expression o ...
ShRNA-specific regulation of FMNL2 expression in P19 cellsYousefLayyous
This video encompasses all the steps and data produced for my graduation project in BSc in Biopharmaceutical science. During the course of the project we modified mammalian cells using Short Hairpin RNA to inhibit the correct function of the cytoskelleton. In this way we studied the importance of FMNL2 for the activation and regulation of actin fibers. Among the methods used are Flourescent microscopy, mamallian cell culture, cloning and flow cytometry.
Similar to Differential Gene Expression Analysis of RNA Seq and Microarray Data highlights role of Hfq gene in expression regulation and parthenogenesis (20)
The dimensions of healthcare quality refer to various attributes or aspects that define the standard of healthcare services. These dimensions are used to evaluate, measure, and improve the quality of care provided to patients. A comprehensive understanding of these dimensions ensures that healthcare systems can address various aspects of patient care effectively and holistically. Dimensions of Healthcare Quality and Performance of care include the following; Appropriateness, Availability, Competence, Continuity, Effectiveness, Efficiency, Efficacy, Prevention, Respect and Care, Safety as well as Timeliness.
Leading the Way in Nephrology: Dr. David Greene's Work with Stem Cells for Ki...Dr. David Greene Arizona
As we watch Dr. Greene's continued efforts and research in Arizona, it's clear that stem cell therapy holds a promising key to unlocking new doors in the treatment of kidney disease. With each study and trial, we step closer to a world where kidney disease is no longer a life sentence but a treatable condition, thanks to pioneers like Dr. David Greene.
Medical Technology Tackles New Health Care Demand - Research Report - March 2...pchutichetpong
M Capital Group (“MCG”) predicts that with, against, despite, and even without the global pandemic, the medical technology (MedTech) industry shows signs of continuous healthy growth, driven by smaller, faster, and cheaper devices, growing demand for home-based applications, technological innovation, strategic acquisitions, investments, and SPAC listings. MCG predicts that this should reflects itself in annual growth of over 6%, well beyond 2028.
According to Chris Mouchabhani, Managing Partner at M Capital Group, “Despite all economic scenarios that one may consider, beyond overall economic shocks, medical technology should remain one of the most promising and robust sectors over the short to medium term and well beyond 2028.”
There is a movement towards home-based care for the elderly, next generation scanning and MRI devices, wearable technology, artificial intelligence incorporation, and online connectivity. Experts also see a focus on predictive, preventive, personalized, participatory, and precision medicine, with rising levels of integration of home care and technological innovation.
The average cost of treatment has been rising across the board, creating additional financial burdens to governments, healthcare providers and insurance companies. According to MCG, cost-per-inpatient-stay in the United States alone rose on average annually by over 13% between 2014 to 2021, leading MedTech to focus research efforts on optimized medical equipment at lower price points, whilst emphasizing portability and ease of use. Namely, 46% of the 1,008 medical technology companies in the 2021 MedTech Innovator (“MTI”) database are focusing on prevention, wellness, detection, or diagnosis, signaling a clear push for preventive care to also tackle costs.
In addition, there has also been a lasting impact on consumer and medical demand for home care, supported by the pandemic. Lockdowns, closure of care facilities, and healthcare systems subjected to capacity pressure, accelerated demand away from traditional inpatient care. Now, outpatient care solutions are driving industry production, with nearly 70% of recent diagnostics start-up companies producing products in areas such as ambulatory clinics, at-home care, and self-administered diagnostics.
CHAPTER 1 SEMESTER V PREVENTIVE-PEDIATRICS.pdfSachin Sharma
This content provides an overview of preventive pediatrics. It defines preventive pediatrics as preventing disease and promoting children's physical, mental, and social well-being to achieve positive health. It discusses antenatal, postnatal, and social preventive pediatrics. It also covers various child health programs like immunization, breastfeeding, ICDS, and the roles of organizations like WHO, UNICEF, and nurses in preventive pediatrics.
R3 Stem Cells and Kidney Repair A New Horizon in Nephrology.pptxR3 Stem Cell
R3 Stem Cells and Kidney Repair: A New Horizon in Nephrology" explores groundbreaking advancements in the use of R3 stem cells for kidney disease treatment. This insightful piece delves into the potential of these cells to regenerate damaged kidney tissue, offering new hope for patients and reshaping the future of nephrology.
QA Paediatric dentistry department, Hospital Melaka 2020Azreen Aj
QA study - To improve the 6th monthly recall rate post-comprehensive dental treatment under general anaesthesia in paediatric dentistry department, Hospital Melaka
Global launch of the Healthy Ageing and Prevention Index 2nd wave – alongside...ILC- UK
The Healthy Ageing and Prevention Index is an online tool created by ILC that ranks countries on six metrics including, life span, health span, work span, income, environmental performance, and happiness. The Index helps us understand how well countries have adapted to longevity and inform decision makers on what must be done to maximise the economic benefits that comes with living well for longer.
Alongside the 77th World Health Assembly in Geneva on 28 May 2024, we launched the second version of our Index, allowing us to track progress and give new insights into what needs to be done to keep populations healthier for longer.
The speakers included:
Professor Orazio Schillaci, Minister of Health, Italy
Dr Hans Groth, Chairman of the Board, World Demographic & Ageing Forum
Professor Ilona Kickbusch, Founder and Chair, Global Health Centre, Geneva Graduate Institute and co-chair, World Health Summit Council
Dr Natasha Azzopardi Muscat, Director, Country Health Policies and Systems Division, World Health Organisation EURO
Dr Marta Lomazzi, Executive Manager, World Federation of Public Health Associations
Dr Shyam Bishen, Head, Centre for Health and Healthcare and Member of the Executive Committee, World Economic Forum
Dr Karin Tegmark Wisell, Director General, Public Health Agency of Sweden
CHAPTER 1 SEMESTER V - ROLE OF PEADIATRIC NURSE.pdfSachin Sharma
Pediatric nurses play a vital role in the health and well-being of children. Their responsibilities are wide-ranging, and their objectives can be categorized into several key areas:
1. Direct Patient Care:
Objective: Provide comprehensive and compassionate care to infants, children, and adolescents in various healthcare settings (hospitals, clinics, etc.).
This includes tasks like:
Monitoring vital signs and physical condition.
Administering medications and treatments.
Performing procedures as directed by doctors.
Assisting with daily living activities (bathing, feeding).
Providing emotional support and pain management.
2. Health Promotion and Education:
Objective: Promote healthy behaviors and educate children, families, and communities about preventive healthcare.
This includes tasks like:
Administering vaccinations.
Providing education on nutrition, hygiene, and development.
Offering breastfeeding and childbirth support.
Counseling families on safety and injury prevention.
3. Collaboration and Advocacy:
Objective: Collaborate effectively with doctors, social workers, therapists, and other healthcare professionals to ensure coordinated care for children.
Objective: Advocate for the rights and best interests of their patients, especially when children cannot speak for themselves.
This includes tasks like:
Communicating effectively with healthcare teams.
Identifying and addressing potential risks to child welfare.
Educating families about their child's condition and treatment options.
4. Professional Development and Research:
Objective: Stay up-to-date on the latest advancements in pediatric healthcare through continuing education and research.
Objective: Contribute to improving the quality of care for children by participating in research initiatives.
This includes tasks like:
Attending workshops and conferences on pediatric nursing.
Participating in clinical trials related to child health.
Implementing evidence-based practices into their daily routines.
By fulfilling these objectives, pediatric nurses play a crucial role in ensuring the optimal health and well-being of children throughout all stages of their development.
Artificial Intelligence to Optimize Cardiovascular Therapy
Differential Gene Expression Analysis of RNA Seq and Microarray Data highlights role of Hfq gene in expression regulation and parthenogenesis
1. Differential Gene Expression Analysis of RNA Seq and Microarray Data
highlights role of Hfq gene in expression regulation and parthenogenesis
Hfq is a conserved RNA-binding protein with multiple regulatory roles within the prokaryotic
cell. One of the key roles is to promote stable duplex formation between small RNAs and
mRNAs and thus Hfq deletion mutants have pleiotropic phenotypes. This study represents
first attempt to characterize Hfq regulon in bacterial pathogen using an integrative omics
approach by comparing RNA Seq and microarray technique.
Source: https://doi.org/10.1073/pnas.1004435107
Hfq protein in action. Figure shows binding mechanism by forming duplex formation.
This recent study by Dienstbier et al (1) performs a comparison across next generation
sequencing using RNA Seq analysis and DNA microarray to establish regulatory action for
Hfq protein in B. pertussis.
Total RNA isolated from biological triplicates of B. pertussis Tohama I strain and its isogenic
hfq strain cultures were analyzed by RNA-seq. Differential gene expression analysis showed
significant modulation of B. pertussis genes including non-coding RNAs. A parallel DNA
microarray analysis was performed and results were compared to determine sensitivity
between the two approaches. Compared to microarray profiling, RNA seq analysis showed
significant sensitivity in retrieving novel genes. RNA Seq analysis pipeline was used for
clustering and principle component analysis.
Heatmap showing clustering of deferentially expressed genes. Differential gene expression
analysis identified 653 significantly modulated B. pertussis genes including 40 non-coding
RNAs and 11 transfer RNA genes.
Gene Ontology (GO) analysis provided insights about the functional profiles of
Hfq-dependent genes. As displayed in figure below, genes related to Translation, Regulation
2. of transcription, and Transmembrane transport terms were up regulated while genes
belonging to Transmembrane transport, Iron–sulfur cluster assembly, Oxido-reduction
process, Pathogenesis, and Protein secretion by the type III secretion system terms were
enriched among the transcripts which were significantly down regulated in the hfq mutant.
Gene Ontology (GO) analysis for functional profiling
Above study clearly highlights advantages of next generation sequencing - Transcriptome
sequencing or RNA seq and bioinformatics RNA Seq analysis pipelines to establish
differential gene expression profiles.
(1) Comparative Integrated Omics Analysis of the Hfq Regulon in Bordetella pertussis.
Int J Mol Sci. 2019 Jun 24;20(12). pii: E3073. doi: 10.3390/ijms20123073.