- The human bB1-crystallin promoter can efficiently drive expression of foreign proteins like GFP in the zebrafish lens, providing a way to introduce crystallins to study prevention of cataract in the cloche mutant.
- The zebrafish aBa- and aBb-crystallin promoters did not drive expression in the embryonic lens, suggesting their roles change during development. Neither will be useful for introducing proteins to the lens.
- Injecting a morpholino to block aA-crystallin caused cataracts in embryos heterozygous for the cloche mutation but not in wildtype, indicating the cloche lens is more susceptible to cataract formation due to
1. Arf6, but not Arf1, is enriched at the fusogenic synapse in Drosophila embryos where myoblast fusion occurs.
2. Myoblast fusion is not affected in arf6 maternal/zygotic mutant embryos, suggesting redundancy with other Arf proteins.
3. Analysis of dominant negative and constitutively active Arf1 and Arf6 mutants found that Arf1 plays an important role in myoblast fusion, while Arf6 may function redundantly with other Arfs.
The document summarizes research identifying genes required for cytoplasmic localization in early C. elegans embryos. Key findings include:
1) Mutations in maternal effect genes disrupted cytoplasmic localization, leading to defects like equal first cleavage, altered second cleavages, abnormal P granule localization, abnormal cell differentiation, and sterility.
2) Genes identified included par-1, par-2, par-3, and par-4. Mutations caused mispositioning of spindles, altered cleavage timing, and improper P granule localization.
3) The par genes appear to function in a common process governing cleavage patterning, intestinal differentiation, and P granule localization, and their mutation
Zinc finger nucleases (ZFNs) are artificial proteins used to edit genes. They contain zinc finger peptides that recognize DNA sequences and nuclease domains that cut DNA. The objective of this study was to generate mastitis-resistant cows using ZFNs. Researchers used ZFNs to insert the human lysozyme gene into the beta-casein locus of bovine cells. The modified cells were used in nuclear transfer to produce cloned cows that secreted human lysozyme in their milk, enhancing resistance to bacterial infection. The results demonstrated that ZFNs can successfully and precisely edit the bovine genome to produce transgenic livestock with improved traits.
This study investigated whether co-exposure to ellagic acid (EA) could mitigate the adverse effects of aflatoxin B1 (AFB1) on visual development in zebrafish larvae. Larvae were exposed to AFB1 alone or with low, medium, and high doses of EA. Visual ability was assessed using optokinetic response testing. While AFB1 alone did not significantly impair vision, co-exposure to medium and high EA doses with AFB1 reduced visual responses more than AFB1 alone. Additionally, higher EA vehicle controls alone impaired vision, suggesting issues with the EA exposure method. In conclusion, co-exposure to EA did not protect and may have exacerbated the effects of AFB
The document discusses mechanisms that promote mono-orientation of sister kinetochores during meiosis I in yeast. It finds that Zip1, a protein involved in synaptonemal complex formation, interacts with components of the Ndc80 kinetochore complex and delays bi-orientation of sister kinetochores when expressed in mitosis. This suggests Zip1 may regulate mono-orientation of sisters similarly to how it arranges chromosomes during synapsis. Deletion of ZIP1 in mutants disrupts this mono-orientation, indicating Zip1 prevents bi-orientation through interacting with functional kinetochores.
1. Adult stem cells isolated from fat (h-ASC) have shown promising results in treating various diseases and improving health. They have been tested for over 9 years in canine clinics with remarkable results and early human trials show benefits for conditions like heart and lung disease.
2. h-ASC are abundant, inexpensive, and simple to produce autologously via liposuction. They may provide hope for currently untreatable diseases and have anti-aging properties. NASA is studying their potential for treating astronaut health issues in space.
3. However, legal and regulatory issues regarding stem cells and their classification have limited clinical availability in the US despite their autologous nature and positive results in other countries. Further research
1) The objective of the laboratory was to transform E. coli bacteria with a gene from jellyfish that codes for Green Fluorescent Protein (GFP).
2) The transformation used a CaCI2 solution and the bacteria were incubated on agar plates containing the transformed samples.
3) Examination of the plates under UV light showed expression of GFP in some samples, and SDS-PAGE electrophoresis identified the GFP protein band at 27 kD in samples run on a polyacrylamide gel.
The document discusses mechanisms that promote mono-orientation of sister kinetochores during meiosis I. It reports that Zip1, a protein involved in synaptonemal complex formation, interacts with components of the Ndc80 kinetochore complex and its deletion reduces mono-orientation. Experiments show that Zip1 expression in mitosis delays bi-orientation of sisters and influences kinetochore orientation in meiosis I mutants. The results suggest Zip1 may prevent bi-orientation of functional sister kinetochores during meiosis I.
1. Arf6, but not Arf1, is enriched at the fusogenic synapse in Drosophila embryos where myoblast fusion occurs.
2. Myoblast fusion is not affected in arf6 maternal/zygotic mutant embryos, suggesting redundancy with other Arf proteins.
3. Analysis of dominant negative and constitutively active Arf1 and Arf6 mutants found that Arf1 plays an important role in myoblast fusion, while Arf6 may function redundantly with other Arfs.
The document summarizes research identifying genes required for cytoplasmic localization in early C. elegans embryos. Key findings include:
1) Mutations in maternal effect genes disrupted cytoplasmic localization, leading to defects like equal first cleavage, altered second cleavages, abnormal P granule localization, abnormal cell differentiation, and sterility.
2) Genes identified included par-1, par-2, par-3, and par-4. Mutations caused mispositioning of spindles, altered cleavage timing, and improper P granule localization.
3) The par genes appear to function in a common process governing cleavage patterning, intestinal differentiation, and P granule localization, and their mutation
Zinc finger nucleases (ZFNs) are artificial proteins used to edit genes. They contain zinc finger peptides that recognize DNA sequences and nuclease domains that cut DNA. The objective of this study was to generate mastitis-resistant cows using ZFNs. Researchers used ZFNs to insert the human lysozyme gene into the beta-casein locus of bovine cells. The modified cells were used in nuclear transfer to produce cloned cows that secreted human lysozyme in their milk, enhancing resistance to bacterial infection. The results demonstrated that ZFNs can successfully and precisely edit the bovine genome to produce transgenic livestock with improved traits.
This study investigated whether co-exposure to ellagic acid (EA) could mitigate the adverse effects of aflatoxin B1 (AFB1) on visual development in zebrafish larvae. Larvae were exposed to AFB1 alone or with low, medium, and high doses of EA. Visual ability was assessed using optokinetic response testing. While AFB1 alone did not significantly impair vision, co-exposure to medium and high EA doses with AFB1 reduced visual responses more than AFB1 alone. Additionally, higher EA vehicle controls alone impaired vision, suggesting issues with the EA exposure method. In conclusion, co-exposure to EA did not protect and may have exacerbated the effects of AFB
The document discusses mechanisms that promote mono-orientation of sister kinetochores during meiosis I in yeast. It finds that Zip1, a protein involved in synaptonemal complex formation, interacts with components of the Ndc80 kinetochore complex and delays bi-orientation of sister kinetochores when expressed in mitosis. This suggests Zip1 may regulate mono-orientation of sisters similarly to how it arranges chromosomes during synapsis. Deletion of ZIP1 in mutants disrupts this mono-orientation, indicating Zip1 prevents bi-orientation through interacting with functional kinetochores.
1. Adult stem cells isolated from fat (h-ASC) have shown promising results in treating various diseases and improving health. They have been tested for over 9 years in canine clinics with remarkable results and early human trials show benefits for conditions like heart and lung disease.
2. h-ASC are abundant, inexpensive, and simple to produce autologously via liposuction. They may provide hope for currently untreatable diseases and have anti-aging properties. NASA is studying their potential for treating astronaut health issues in space.
3. However, legal and regulatory issues regarding stem cells and their classification have limited clinical availability in the US despite their autologous nature and positive results in other countries. Further research
1) The objective of the laboratory was to transform E. coli bacteria with a gene from jellyfish that codes for Green Fluorescent Protein (GFP).
2) The transformation used a CaCI2 solution and the bacteria were incubated on agar plates containing the transformed samples.
3) Examination of the plates under UV light showed expression of GFP in some samples, and SDS-PAGE electrophoresis identified the GFP protein band at 27 kD in samples run on a polyacrylamide gel.
The document discusses mechanisms that promote mono-orientation of sister kinetochores during meiosis I. It reports that Zip1, a protein involved in synaptonemal complex formation, interacts with components of the Ndc80 kinetochore complex and its deletion reduces mono-orientation. Experiments show that Zip1 expression in mitosis delays bi-orientation of sisters and influences kinetochore orientation in meiosis I mutants. The results suggest Zip1 may prevent bi-orientation of functional sister kinetochores during meiosis I.
The study examined gene expression levels of TGF-β1, IGF-1, and IL-1β in bladder tissue samples from wildtype and cystinuria knockout mice of different ages. RNA was extracted from the samples and converted to cDNA, which was then used to perform qPCR to analyze gene expression levels. The results found no statistically significant differences in TGF-β1 or IGF-1 expression levels between genotypes or age groups. However, one 5-month old knockout mouse sample showed unusually high IGF-1 expression levels in multiple tests, suggesting another gene may be affecting its expression. Further research is needed to verify this possibility and test IL-1β expression levels.
This study investigated the effects of developmental exposure to the organophosphate chlorpyrifos on social play behavior and endocannabinoid system activation in rats. The researchers found that rats exposed to chlorpyrifos engaged in more social play behaviors compared to controls. However, chlorpyrifos exposure did not affect phosphorylation of cannabinoid receptors in brain regions involved in social play, suggesting that increased social play is not related to changes in endocannabinoid system activation. While endocannabinoid signaling may play a role in altered social behavior, chlorpyrifos may induce changes in other neurotransmitter systems like dopamine that are important for social play.
Uncovering novel candidate genes for pyridoxine-responsive epilepsy in a cons...Golden Helix Inc
This document summarizes Hilal Al Shekaili's work on characterizing the genetic cause of pyridoxine-dependent epilepsy (PDE) in an Omani family. [1] Runs of homozygosity mapping and whole-exome sequencing identified two candidate genes involved in vitamin transport and neuropeptide processing. [2] Further studies are planned to validate the candidate genes and recruit additional families. [3] Identifying new PDE genes could improve treatment and fill knowledge gaps in pyridoxine metabolism.
R lh supplementation to rfsh in gnrh antagonist cyclesAlfredo Nazzaro
1) The study compared the effects of recombinant FSH (rFSH) supplementation alone versus rFSH supplemented with recombinant LH (rLH) in GnRH antagonist cycles on implantation and pregnancy rates.
2) They found that the rFSH + rLH group had higher numbers of retrieved and metaphase II oocytes, higher fertilization rates, and more grade I-II embryos compared to the rFSH only group.
3) The rFSH + rLH group also had higher implantation and clinical pregnancy rates. The authors conclude that rLH supplementation improves outcomes likely by enhancing oocyte quality and endometrial receptivity.
This study investigated the effects of knocking down neuroligin genes on planarian regeneration abilities. Planarians were separated into four groups - a control group with an unc22 gene knockdown and three experimental groups with neuroligin 1A/1B, 2, or 1B/2 gene knockdowns. The worms were cut and imaged over 10 days to observe eyespot regeneration and blastema tissue re-pigmentation. Results showed that single neuroligin knockdowns caused 1-2 day delays in regeneration, while the double 1B/2 knockdown caused more severe 4 day delays. This supports the hypothesis that neuroligins are important for synaptic signaling required for normal planarian regeneration.
There are three main types of cloning: 1) DNA cloning which clones genes for uses like protein production and vaccine development, 2) reproductive cloning which produces genetically identical animals but risks are high inefficiency and potential harm, and 3) therapeutic cloning which produces stem cells for research on treating diseases but risks include killing embryos.
This study examined the effects of inhibiting the Hippo signaling pathway during sea urchin embryogenesis. Researchers treated sea urchin embryos with verteporfin, an inhibitor of YAP which is a key component of the Hippo pathway. They found that inhibiting the Hippo pathway delayed cell cleavage and gastrulation. While marker genes for specific cell types were still expressed, the patterns of expression were altered. Inhibition of the Hippo pathway resulted in epithelial cells that were larger in size and fewer in number, suggesting YAP is involved in epithelial organization and proliferation during sea urchin development.
This document summarizes a study that examined the effects of bisphenol A (BPA), bisphenol F (BPF), and high levels of estrogen on oocyte maturation and quality. Mouse oocytes were exposed to varying concentrations of BPA, BPF, and estrogen. The oocytes were then analyzed for spindle formation, centrosome position, and chromosome alignment. Oocytes exposed to BPA, BPF, and high estrogen showed lower maturation rates and more chromosome abnormalities compared to the control group. BPA exposure resulted in more chromosomal errors than BPF exposure. The findings suggest that exposure to synthetic estrogens like BPA and BPF can negatively impact oocyte development and potentially cause birth defects.
Effects of knockout of antioxidant genes on spermatogenesisxsonixs
1) The study examined the effects of knocking out antioxidant genes Gclm and Nrf2 on sperm production and morphology in mice.
2) For Nrf2 knockout mice, results showed significantly decreased testicular and epididymal sperm counts compared to wild type mice, but no significant effects on sperm morphology.
3) For Gclm knockout mice, results found no significant effects on sperm morphology but a trend of higher percentages of immature sperm compared to wild type mice.
Analysis of sypK gene in Vibrio fischeri BiofilmsAishwarya Raj
The document analyzes the sypK gene in Vibrio fischeri biofilms. The sypK gene codes for a putative oligosaccharide translocase that promotes bacterial motility and biofilm formation. The purpose of the investigation was to determine if these two phenotypes, motility and biofilm formation, can be expressed independently by the bacterium and to identify essential amino acids in sypK. Through generating mutant strains, evidence suggests that different amino acids are required for biofilm formation versus motility as some mutants exhibited motility without biofilm formation. However, DNA sequencing found no significant mutations, so it was theorized that expression level of sypK, rather than sequence, determines the phenotypes expressed.
Caspase activation contributes to astrogliosis without inducing apoptosis in astrocytes. The study found that treating cultured neonatal rat astrocytes with dibutryl cAMP or beta-amyloid peptide, stimuli known to induce astrogliosis, led to increased caspase activity and expression of active caspase-3 without cell death. Inhibition of caspases attenuated the increased expression of glutamine synthetase and fibroblast growth factor-2, markers of astrogliosis. The results suggest caspases play a non-apoptotic role in regulating astrogliosis in astrocytes following brain injury.
The document discusses the green fluorescent protein (GFP) discovered in the jellyfish Aequorea victoria. GFP causes the jellyfish to glow green through bioluminescence. In the 1960s, Osamu Shimomura isolated GFP from Aequorea victoria and his work helped uncover how GFP fluorescence works. GFP has since become an important tool in molecular and cell biology by allowing researchers to track cellular movements and protein expressions through genetic tagging with the GFP gene.
This document describes a study that investigated the seminal plasma proteome of Holstein bulls with low and high sperm freezability. Label-free mass spectrometry identified 1,445 seminal plasma proteins. There were 338 proteins differentially expressed between bulls with low and high freezability. Specific proteins were identified as markers of each phenotype, with BSP5 and seminal ribonuclease associated with high freezability and spermadhesin-1, gelsolin, and peroxiredoxin-5 associated with low freezability. Regression models found sperm freezability scores were related to levels of peroxiredoxin-5, spermadhesin-1, and their interaction. This research provides insights
GBA activity was assessed in cell lines with and without GBA knockout. Activity decreased in a dose-dependent manner with CBE treatment but not ambroxol or isofagomine after short incubation. Mutant B lymphocytes and fibroblasts generally had lower GBA levels and activity than controls, with some donor variability. Treatment of A53T transgenic mouse neurons with ambroxol and isofagomine decreased α-synuclein levels, indicating potential as chaperones to increase GBA activity and reduce α-synuclein aggregates.
This document summarizes a study examining the role of the Brg1 gene during early embryonic development in mice. The researchers conditionally deleted the Brg1 gene beginning at gastrulation using a tamoxifen-inducible Cre system. They found that Brg1 deficiency resulted in increased apoptosis, growth retardation, and embryonic death. Gene expression analysis revealed upregulation of genes that negatively regulate cell growth and cell cycle progression. Additionally, the p53 protein accumulated in Brg1-deficient embryos and activated p53-dependent pathways. The researchers provide evidence that Brg1 functions in part via regulating the p53 program to constrain gene expression and facilitate rapid embryonic growth.
Modulation of pluripotency in the porcine embryo and i ps cells (Dec.27,2012) Ahmad Usama
1) The study investigated signaling pathways involved in formation of the porcine inner cell mass (ICM) and methods to increase pluripotency in porcine embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs).
2) They found that inhibiting MEK and GSK3β signaling pathways in porcine embryos and iPSCs, combined with LIF treatment, promoted a naive pluripotent state similar to mouse ESCs.
3) Naive piPSCs expressed genes associated with naive pluripotency and showed higher germline differentiation potential than non-naive piPSCs. The study provides insights into regulating pluripotency in
The document describes a pilot study that investigated the presence of DNA in blastocyst fluids (BFs) and whether the chromosomal status predicted by analyzing this DNA corresponds to the status in trophectoderm (TE) cells and the whole embryo. The study found that:
1) DNA was detected in the BFs of 76.5% of blastocysts tested, allowing chromosomal analysis of these samples.
2) In 97.4% of cases, the ploidy condition (euploid vs. aneuploid) predicted by BF analysis matched the condition in TE cells.
3) BF analysis predicted the ploidy condition of the whole embryo with 100% accuracy
Gfp application in bacterial dynamics and disease diagnosisgarima shrinet
GFP has numerous applications for visualizing bacterial dynamics and disease diagnosis. It can be used to track gene expression, protein localization, and cell processes in real-time without disrupting bacterial viability. Fusion of GFP to proteins of interest allows visualization of processes like cell division, sporulation, and biofilm formation. GFP reporters also help identify genes expressed during bacterial infection and pathogenesis. Advances like FRET biosensors now enable monitoring of intracellular calcium and protein-protein interactions. Overall, GFP has revolutionized the study of bacterial physiology, gene regulation, and host-pathogen interactions.
MAIN MOLECULAR MARKERS OF ORAL SQUAMOUS CELL CARCINOMA / dental implant cour...Indian dental academy
The Indian Dental Academy is the Leader in continuing dental education , training dentists in all aspects of dentistry and
offering a wide range of dental certified courses in different formats.for more details please visit
www.indiandentalacademy.com
This document discusses materials and methods used in a study involving the chemical fipronil and zinc. Twenty male albino rats were divided into four groups of five rats each: a control group, a zinc group that received zinc supplementation, a fipronil group exposed to the insecticide fipronil, and a combination group exposed to both zinc and fipronil. Biochemical assays were conducted to assess oxidative stress markers like superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase, glutathione, lipid peroxidation, and total protein in the rats. Chemicals used including fipronil and zinc sulfate were obtained from reputable suppliers. Kits for the biochemical assays were purchased from a diagnostic
This study examined the effects of ethanol exposure on early eye development in chicken embryos. Chicken eggs were injected with varying concentrations of ethanol or saline on embryonic day 1, then harvested after 40 hours or 5 days. Ethanol exposure caused dose-dependent abnormalities including microphthalmia, growth retardation, and increased vascularization. Expression of genes Bmp4 and Pax2, important for eye development, was reduced in a dose-dependent manner with ethanol exposure. The results suggest ethanol disrupts key molecular signals like Bmp4 and Pax2 that govern early eye formation, potentially explaining ocular defects seen in fetal alcohol syndrome.
The document discusses factors affecting fertilization using intracytoplasmic sperm injection (ICSI). It outlines sperm-related factors like abnormal morphology, DNA damage, and methods to select viable immotile sperm. It also discusses oocyte-related factors like abnormal morphology and maturation stages. The document emphasizes that low or no fertilization can occur due to few mature oocytes, oocyte activation failure, or lack of appropriate sperm, but repeated ICSI attempts can achieve fertilization in 85% of cases.
The study examined gene expression levels of TGF-β1, IGF-1, and IL-1β in bladder tissue samples from wildtype and cystinuria knockout mice of different ages. RNA was extracted from the samples and converted to cDNA, which was then used to perform qPCR to analyze gene expression levels. The results found no statistically significant differences in TGF-β1 or IGF-1 expression levels between genotypes or age groups. However, one 5-month old knockout mouse sample showed unusually high IGF-1 expression levels in multiple tests, suggesting another gene may be affecting its expression. Further research is needed to verify this possibility and test IL-1β expression levels.
This study investigated the effects of developmental exposure to the organophosphate chlorpyrifos on social play behavior and endocannabinoid system activation in rats. The researchers found that rats exposed to chlorpyrifos engaged in more social play behaviors compared to controls. However, chlorpyrifos exposure did not affect phosphorylation of cannabinoid receptors in brain regions involved in social play, suggesting that increased social play is not related to changes in endocannabinoid system activation. While endocannabinoid signaling may play a role in altered social behavior, chlorpyrifos may induce changes in other neurotransmitter systems like dopamine that are important for social play.
Uncovering novel candidate genes for pyridoxine-responsive epilepsy in a cons...Golden Helix Inc
This document summarizes Hilal Al Shekaili's work on characterizing the genetic cause of pyridoxine-dependent epilepsy (PDE) in an Omani family. [1] Runs of homozygosity mapping and whole-exome sequencing identified two candidate genes involved in vitamin transport and neuropeptide processing. [2] Further studies are planned to validate the candidate genes and recruit additional families. [3] Identifying new PDE genes could improve treatment and fill knowledge gaps in pyridoxine metabolism.
R lh supplementation to rfsh in gnrh antagonist cyclesAlfredo Nazzaro
1) The study compared the effects of recombinant FSH (rFSH) supplementation alone versus rFSH supplemented with recombinant LH (rLH) in GnRH antagonist cycles on implantation and pregnancy rates.
2) They found that the rFSH + rLH group had higher numbers of retrieved and metaphase II oocytes, higher fertilization rates, and more grade I-II embryos compared to the rFSH only group.
3) The rFSH + rLH group also had higher implantation and clinical pregnancy rates. The authors conclude that rLH supplementation improves outcomes likely by enhancing oocyte quality and endometrial receptivity.
This study investigated the effects of knocking down neuroligin genes on planarian regeneration abilities. Planarians were separated into four groups - a control group with an unc22 gene knockdown and three experimental groups with neuroligin 1A/1B, 2, or 1B/2 gene knockdowns. The worms were cut and imaged over 10 days to observe eyespot regeneration and blastema tissue re-pigmentation. Results showed that single neuroligin knockdowns caused 1-2 day delays in regeneration, while the double 1B/2 knockdown caused more severe 4 day delays. This supports the hypothesis that neuroligins are important for synaptic signaling required for normal planarian regeneration.
There are three main types of cloning: 1) DNA cloning which clones genes for uses like protein production and vaccine development, 2) reproductive cloning which produces genetically identical animals but risks are high inefficiency and potential harm, and 3) therapeutic cloning which produces stem cells for research on treating diseases but risks include killing embryos.
This study examined the effects of inhibiting the Hippo signaling pathway during sea urchin embryogenesis. Researchers treated sea urchin embryos with verteporfin, an inhibitor of YAP which is a key component of the Hippo pathway. They found that inhibiting the Hippo pathway delayed cell cleavage and gastrulation. While marker genes for specific cell types were still expressed, the patterns of expression were altered. Inhibition of the Hippo pathway resulted in epithelial cells that were larger in size and fewer in number, suggesting YAP is involved in epithelial organization and proliferation during sea urchin development.
This document summarizes a study that examined the effects of bisphenol A (BPA), bisphenol F (BPF), and high levels of estrogen on oocyte maturation and quality. Mouse oocytes were exposed to varying concentrations of BPA, BPF, and estrogen. The oocytes were then analyzed for spindle formation, centrosome position, and chromosome alignment. Oocytes exposed to BPA, BPF, and high estrogen showed lower maturation rates and more chromosome abnormalities compared to the control group. BPA exposure resulted in more chromosomal errors than BPF exposure. The findings suggest that exposure to synthetic estrogens like BPA and BPF can negatively impact oocyte development and potentially cause birth defects.
Effects of knockout of antioxidant genes on spermatogenesisxsonixs
1) The study examined the effects of knocking out antioxidant genes Gclm and Nrf2 on sperm production and morphology in mice.
2) For Nrf2 knockout mice, results showed significantly decreased testicular and epididymal sperm counts compared to wild type mice, but no significant effects on sperm morphology.
3) For Gclm knockout mice, results found no significant effects on sperm morphology but a trend of higher percentages of immature sperm compared to wild type mice.
Analysis of sypK gene in Vibrio fischeri BiofilmsAishwarya Raj
The document analyzes the sypK gene in Vibrio fischeri biofilms. The sypK gene codes for a putative oligosaccharide translocase that promotes bacterial motility and biofilm formation. The purpose of the investigation was to determine if these two phenotypes, motility and biofilm formation, can be expressed independently by the bacterium and to identify essential amino acids in sypK. Through generating mutant strains, evidence suggests that different amino acids are required for biofilm formation versus motility as some mutants exhibited motility without biofilm formation. However, DNA sequencing found no significant mutations, so it was theorized that expression level of sypK, rather than sequence, determines the phenotypes expressed.
Caspase activation contributes to astrogliosis without inducing apoptosis in astrocytes. The study found that treating cultured neonatal rat astrocytes with dibutryl cAMP or beta-amyloid peptide, stimuli known to induce astrogliosis, led to increased caspase activity and expression of active caspase-3 without cell death. Inhibition of caspases attenuated the increased expression of glutamine synthetase and fibroblast growth factor-2, markers of astrogliosis. The results suggest caspases play a non-apoptotic role in regulating astrogliosis in astrocytes following brain injury.
The document discusses the green fluorescent protein (GFP) discovered in the jellyfish Aequorea victoria. GFP causes the jellyfish to glow green through bioluminescence. In the 1960s, Osamu Shimomura isolated GFP from Aequorea victoria and his work helped uncover how GFP fluorescence works. GFP has since become an important tool in molecular and cell biology by allowing researchers to track cellular movements and protein expressions through genetic tagging with the GFP gene.
This document describes a study that investigated the seminal plasma proteome of Holstein bulls with low and high sperm freezability. Label-free mass spectrometry identified 1,445 seminal plasma proteins. There were 338 proteins differentially expressed between bulls with low and high freezability. Specific proteins were identified as markers of each phenotype, with BSP5 and seminal ribonuclease associated with high freezability and spermadhesin-1, gelsolin, and peroxiredoxin-5 associated with low freezability. Regression models found sperm freezability scores were related to levels of peroxiredoxin-5, spermadhesin-1, and their interaction. This research provides insights
GBA activity was assessed in cell lines with and without GBA knockout. Activity decreased in a dose-dependent manner with CBE treatment but not ambroxol or isofagomine after short incubation. Mutant B lymphocytes and fibroblasts generally had lower GBA levels and activity than controls, with some donor variability. Treatment of A53T transgenic mouse neurons with ambroxol and isofagomine decreased α-synuclein levels, indicating potential as chaperones to increase GBA activity and reduce α-synuclein aggregates.
This document summarizes a study examining the role of the Brg1 gene during early embryonic development in mice. The researchers conditionally deleted the Brg1 gene beginning at gastrulation using a tamoxifen-inducible Cre system. They found that Brg1 deficiency resulted in increased apoptosis, growth retardation, and embryonic death. Gene expression analysis revealed upregulation of genes that negatively regulate cell growth and cell cycle progression. Additionally, the p53 protein accumulated in Brg1-deficient embryos and activated p53-dependent pathways. The researchers provide evidence that Brg1 functions in part via regulating the p53 program to constrain gene expression and facilitate rapid embryonic growth.
Modulation of pluripotency in the porcine embryo and i ps cells (Dec.27,2012) Ahmad Usama
1) The study investigated signaling pathways involved in formation of the porcine inner cell mass (ICM) and methods to increase pluripotency in porcine embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs).
2) They found that inhibiting MEK and GSK3β signaling pathways in porcine embryos and iPSCs, combined with LIF treatment, promoted a naive pluripotent state similar to mouse ESCs.
3) Naive piPSCs expressed genes associated with naive pluripotency and showed higher germline differentiation potential than non-naive piPSCs. The study provides insights into regulating pluripotency in
The document describes a pilot study that investigated the presence of DNA in blastocyst fluids (BFs) and whether the chromosomal status predicted by analyzing this DNA corresponds to the status in trophectoderm (TE) cells and the whole embryo. The study found that:
1) DNA was detected in the BFs of 76.5% of blastocysts tested, allowing chromosomal analysis of these samples.
2) In 97.4% of cases, the ploidy condition (euploid vs. aneuploid) predicted by BF analysis matched the condition in TE cells.
3) BF analysis predicted the ploidy condition of the whole embryo with 100% accuracy
Gfp application in bacterial dynamics and disease diagnosisgarima shrinet
GFP has numerous applications for visualizing bacterial dynamics and disease diagnosis. It can be used to track gene expression, protein localization, and cell processes in real-time without disrupting bacterial viability. Fusion of GFP to proteins of interest allows visualization of processes like cell division, sporulation, and biofilm formation. GFP reporters also help identify genes expressed during bacterial infection and pathogenesis. Advances like FRET biosensors now enable monitoring of intracellular calcium and protein-protein interactions. Overall, GFP has revolutionized the study of bacterial physiology, gene regulation, and host-pathogen interactions.
MAIN MOLECULAR MARKERS OF ORAL SQUAMOUS CELL CARCINOMA / dental implant cour...Indian dental academy
The Indian Dental Academy is the Leader in continuing dental education , training dentists in all aspects of dentistry and
offering a wide range of dental certified courses in different formats.for more details please visit
www.indiandentalacademy.com
This document discusses materials and methods used in a study involving the chemical fipronil and zinc. Twenty male albino rats were divided into four groups of five rats each: a control group, a zinc group that received zinc supplementation, a fipronil group exposed to the insecticide fipronil, and a combination group exposed to both zinc and fipronil. Biochemical assays were conducted to assess oxidative stress markers like superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase, glutathione, lipid peroxidation, and total protein in the rats. Chemicals used including fipronil and zinc sulfate were obtained from reputable suppliers. Kits for the biochemical assays were purchased from a diagnostic
This study examined the effects of ethanol exposure on early eye development in chicken embryos. Chicken eggs were injected with varying concentrations of ethanol or saline on embryonic day 1, then harvested after 40 hours or 5 days. Ethanol exposure caused dose-dependent abnormalities including microphthalmia, growth retardation, and increased vascularization. Expression of genes Bmp4 and Pax2, important for eye development, was reduced in a dose-dependent manner with ethanol exposure. The results suggest ethanol disrupts key molecular signals like Bmp4 and Pax2 that govern early eye formation, potentially explaining ocular defects seen in fetal alcohol syndrome.
The document discusses factors affecting fertilization using intracytoplasmic sperm injection (ICSI). It outlines sperm-related factors like abnormal morphology, DNA damage, and methods to select viable immotile sperm. It also discusses oocyte-related factors like abnormal morphology and maturation stages. The document emphasizes that low or no fertilization can occur due to few mature oocytes, oocyte activation failure, or lack of appropriate sperm, but repeated ICSI attempts can achieve fertilization in 85% of cases.
UTF-8''Final Assessing post-synaptic partners of Dentate Granule Cells in a M...Grant Pizzo
1) The document describes an experiment assessing how dentate granule cell (DGC) axon terminals and their post-synaptic partners change in a rat model of temporal lobe epilepsy (TLE).
2) The researcher induced seizures in rats using pilocarpine and later injected retroviruses to label dividing DGCs. Tissue was then analyzed to identify post-synaptic partners of DGCs using antibodies targeting interneurons and mossy cells.
3) Preliminary results showed that antibodies for GAD67 and GluR2 successfully labeled the correct cell types, but need further refinement, while an mGluR1/5 antibody did not label interneurons in the dentate gyrus
Effects of prenatal alcohol exposure on the visual system of monkeys measured...BARRY STANLEY 2 fasd
- The document describes a study that used electroretinography (ERG) to examine the effects of prenatal alcohol exposure on retinal development and function in vervet monkeys of different ages.
- ERG responses under dark-adapted (scotopic) and light-adapted (photopic) conditions were measured in 37 monkeys exposed to alcohol prenatally and 41 unexposed control monkeys ranging from 3 to 12 years old.
- Results showed that both age and prenatal alcohol exposure affected ERG responses. Under photopic conditions, amplitudes increased with age and were higher in alcohol-exposed monkeys, related to OFF- and ON-pathways. Under scotopic conditions, amplitudes were decreased in young alcohol
Pnut and Ena may function together during epithelial morphogenesis in Drosophila embryos. A genetic screen identified Pnut as enhancing defects caused by an overactive Abl mutant. Pnut is required for cytokinesis via its role in the actin/myosin contractile ring. The authors found that a mutation in pnut increased dorsal closure and head involution defects in embryos mutant for ena. This suggests Pnut may function with Ena during these processes, possibly through roles in analogous actin/myosin structures that drive morphogenesis. Future experiments will examine Pnut and actin localization and dynamics to elucidate how Pnut and Ena cooperate during epithelial remodeling.
Keloid explant culture: a
model for keloid fibroblasts
isolation and cultivation
based on the biological
differences of its specific
regions
ORIGINAL ARTICLE
Wound Healing
1. Manipulating amounts of the lens protein alpha A-crystallin can alter the development of lens
cataract
Cassie Craig, Dept. of Biology & Environmental Science Program, Ashland University, OH
Mentor: Mason Posner, Ph.D.
Examination of lens crystallin promoters
Adult zebrafish were placed in breeding tanks to collect eggs for injection
experiments. All injections were done with a Harvard Apparatus picoinjector.
Linearized plasmids containing either a promoter for aBa-, aBb- or bB1-
crystallin attached to the GFP gene were injected into single-celled zygotes.
Seventy picograms of each plasmid with 0.2% phenol red as a tracer dye was
injected in a volume of 1 nanoliter. Twenty-four hours post fertilization (hpf)
the chemical PTU was added to the water holding the injected embryos to
block melanin production so that GFP expression could be visualized by
fluorescent microscopy.
Morpholino injections to block aA-crystallin production
Using a similar injection technique as above, 4.5 ng of an MO that blocks aA-
crystallin translation was injected into zebrafish embryos, which were then
examined by Nomarski microscopy to identify cataracts and embryos were
sectioned and stained with hematoxylin and eosin to identify any
abnormalities in lens development.
The purpose of this study was to show that altering aA-crystallin levels can
impact the development of cataracts and that the cloche zebrafish mutant can
be a valuable model for studying this costly visual disease.
BACKGROUND
PURPOSE AND HYPOTHESIS
MATERIALS AND METHODS
DISCUSSION
REFERENCES
Cataracts are the leading cause of human blindness. Zebrafish have
become a valuable model for human diseases because of their similar genetics,
ability to produce many offspring for study, and the ease with which they can be
genetically modified. Alpha A and alpha B-crystallin are small heat shock
proteins that are components of the vertebrate eye lens. Mutations in a-
crystallins are known to cause lens diseases such as cataract. A previous study
showed that low levels of A-crystallin are associated with lens cataract in the
cloche zebrafish mutant (Goishi 2006). In this study we examined how altering
a-crystallin levels affect cataract development. Morpholino anti-sense RNAs
(MO) were injected into wildtype and cloche embryos to block the production
of aA-crystallin protein. We previously showed that prevention of aA-
crystallin production had no effect on the wildtype lens (Posner 2013). Here we
tested whether the cloche lens is more susceptible to cataract development. We
also examined the effectiveness of various lens protein promoters to introduce
crystallins into the cloche zebrafish so that we can study their ability to prevent
cataract. This was done by visualizing the ability of various crystallin
promoters to drive the expression of green fluorescent protein (GFP) in
zebrafish embryos. Our results will lead to future studies that use the cloche
zebrafish as a model to test the ability of native and modified a-crystallins to
prevent lens cataract.
ACKNOWLEDGMENTS
Human bB1-crystallin promoter can be used to add proteins to the cloche
zebrafish lens:
Because the cloche zebrafish mutant develops cataract if it has two copies of
the mutant allelle, this strain has the potential to be used as a model system for
studying cataract development and prevention. We show here that the human bB1-
crystallin promoter can efficiently drive expression of a foreign protein into the
zebrafish lens. We can now use this promoter to introduce various native and
modified a-crystallins to determine whether they can prevent the development of
cataract. The work reported here provides a foundation for these future studies.
Zebrafish aBa- and aBa-crystallin promoters are not active in lens:
Adult zebrafish produce both aB-crystallin proteins in their lens.
Interestingly, the promoters for these genes were not active in the embryonic lens.
This suggests that the expression and function of these two proteins changes during
zebrafish development. Our results also show that neither of these promoters will
be useful for introducing proteins into the zebrafish lens. Interestingly, aBa-
crystallin is only found in the lens of adult zebrafish, but here we show it expressed
in skeletal muscle and notochord of embryos. Future studies will need to address
why and how this protein becomes restricted to the lens later in development. This
topic is relevant to a broader question about how the location of protein expression
can evolve.
MO injection causes cataracts in the heterozygous cloche mutant allele:
We previously showed that the lens of wildtype zebrafish develop normally if
aA-crystallin production is blocked by morpholino injection (MO). The results of
this present study, however, show that embryos heterozygous for the cloche mutant
allele will develop cataract after MO injection. This result suggests that the cloche
lens is more susceptible to cataract than wildtype zebrafish, perhaps due to greater
physiological stress related to the cloche mutation. In the heterozygous cloche
embryo one allele is mutant and the other is not. The lens research community has
been interested in determining the role that aA-crystallin may play in preventing
cataract. Our results here suggest that the impact of aA-crystallin may depend on
the genetic background of the individual being examined.
• Goishi, K., Shimizu, A., Najarro, G., Watanabe, S., Rogers, R., Zon, L. I., & Klagsbrun, M.
(2006). αA-crystallin expression prevents γ-crystallin insolubility and cataract formation in
the zebrafish cloche mutant lens. Development (09501991), 133(13), 13.
• Posner, M., Skiba, J., Brown, M., Liang, J. O., Nussbaum, J., & Prior, H. (2013). Loss of the
small heat shock protein αA-crystallin does not lead to detectable defects in early zebrafish
lens development. Experimental Eye Research, 116227-233. doi:10.1016/j.exer.2013.09.007
• Smith, A. A., Wyatt, K., Vacha, J., Vihtelic, T. S., Zigler Jr, J. S., Wistow, G. J., & Posner, M.
(2006). Gene duplication and separation of functions in αB-crystallin from zebrafish (Danio
rerio). FEBS Journal, 273(3), 481-490. doi:10.1111/j.1742-4658.2005.05080.x
Thank you Mason Posner and Kelly Murray for the help and mentoring of the project. Funding for this project
was provided by the Department of Biology. For additional information, please contact ccraig2@ashland.edu or
mposner@ashland.edu
Figure 2: Embryos showed GFP expression in the notochord, skeletal muscle and geometric shapes (insert).
The proportion of embryos with GFP expression in skeletal muscle and notochord differed between the two
promoters. The aBa promoter was more active in notochord while the aBb promoter was more active in skeletal
muscle. Geometric shapes expressing GFP were identified in the skin of the embryos, but reasons for this expression
and the identity of this tissue are unclear. Lens expression was very rare or absent with these promoters. Error bars
indicate standard error (n = 3 and 2 respectively).
Figure 1: The human bb1 promoter drove GFP expression in the zebrafish lens (insert). Two batches of 40 eggs
each were injected with the bb1/GFP plasmid. Over a course of five days the embryos were checked for
expression. GFP Expression occurred after 48 hours in the lens, increased up until 72 hours and then stabilized
after 72 hours. It was found that 38% of the embryos showed GFP expression. There was no expression anywhere
else outside of the lens.
Figure 3: Sections through zebrafish embryos. Wildtype embryos without lens cataracts showed normal lens
development (A). A heterozygous cloche mutant embryo injected with an MO to block aA-crystallin production
shows abnormal lens development (B). The lens contains “dents” and the central fiber cells have not lost their
nuclei, as normally happens during lens development. Fifty percent of injected cloche embryos had cataract.
0%
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0 48 72 120
PercentofEmbryosWithGFP
Expression
Hours post fertilization
The human bB1-crystallin promoter can drive GFP expression in the zebrafish lens
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Skeletal Muscle Notochord Geometric Shapes Lens
AveragePercentofZebrafishwithGFP
Expression
Location on Fish
Alpha-Ba Promotor Injection
Alpha-Bb Promotor Injection
The aBa and aBb promoters differ in expression and show rare expression in lens
MO injections resulted in cataract for cloche embryos but not in wildtype embryos
Picoinjector
Zygote
Yolk
Embryo
A B
RESULTS