Difference between revisions of "User:Z3417458"
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Our stem cell presentation was based on the article below, it discusses the use of CRISPR-Cas9 to
Our stem cell presentation was based on the article below, it discusses the use of CRISPR-Cas9 to treat cataract disease in mice.
Latest revision as of 08:41, 10 June 2015
- 1 Individual Assessments
- 1.1 LAB 12
- 1.2 LAB 11
- 1.3 LAB 9
- 1.4 Peer Reviews
- 1.5 LAB 6
- 1.6 LAB 5
- 1.7 LAB 3
- 1.8 During the lab
- 1.9 LAB 2
- 1.10 During the lab
- 1.11 Reference
- 1.12 LAB 1
- 2 Lab Attendance
"Identify a paper that uses a technique described in the visit to the Ramaciotti Centre that relates to your Group project topic. Add reference to your page and have a couple sentences describing the related technique."
In the labratory several techniques were discussed one of them included 'Illumina Methylation Assays'. The technique uses the Infinium II which uses a 'Beadchip' technology to determine the genome in human DNA methylation. The methylation levels of specific loci within the genome are identified. In addition the Illumina’s Infinium Methylation Assay give quantitative measures of methylation at single CpG-site level, providing high resolution and allows us to interpret epigenetic changes . The use of the technology allows single nucleotide polymorphisms (SNP) genotyping which shows differences in sequences of genes, whole genome sequencing that can identify the genome sequence of an individual or organism, gene expression and transcriptome profiling and epigenetics .
Our stem cell presentation was based on the article below, it discusses the use of CRISPR-Cas9 to treat cataract disease in mice.
This page has been executed really well the layout is structured well and is cohesive with the topics, subtopics and related images. I like that you an image at the start of the page of a proteoglycan structure its pretty. Form your referencing list it is evident that you've research the topic extensively. The amount of content that your have included so far is really good. However I think some sections might need a little more info and detail, especially at the start and end of the page. Make sure you include an introduction into the page to summaries a basic overview of the topic and what your page will cover.I know generally this is something you do towards the end of the assessment but just ensure you do include it as I've noticed pretty much all the groups have one. Your use of images is good they are nice and vibrant, great work doing your own drawings especially those chemical structures. However I'd consider adding more images on how the proteoglycans are formed, maybe try to use a video for it. I like your use of a variety of formats in the images so your use of table, drawings and flow chart. Your in-text citations are all consistent, I've just noticed some links to pubmed articles, I think this maybe because your still going to add information to those sections. Otherwise just ensure that you do cite the the way you've done with all the others. I like that you've included a glossary at the end of the page its clear and easy to understand. Overall I think you've done exceptionally well so far just a few small adjustment here and there content wise and addition of images.
Keep up the great work everyone, good luck ! :)
I think the layout of the page is done well, with all your constituent topics and subtopics. Although I think you need a lot more information in some of the subtopics especially in structure and function and a little on diseases. Based on your reference list it seems that you've done quite a bit of research, you just need to try to add more content and maybe more detail. Your in-text citations are formatted really well, but I think you don't know how to reference the websites so here are some tips. Go to the tab on the left hand side of the main page and click on editing basics, there should be a tab on project referencing. If your referencing a website add these symbols without the brackets and lines . I also noticed on your reference list you have some references that show up on the list and there's an empty space, maybe look over your formula again or refer to the project referencing tab. I like your use of images they are nice and vibrant, good use of a table in the future research topic. Also you've provided a great variety of different formats to show the function and structure of intergrins. This is really great !! I like that you've used a video its wonderful as sometimes content can be difficult and boring to understand. Thus this video is short, clear and a good way to show the interaction with the extracellular matrix. I'd suggest adding more images to the diseases component of the page, if you have trouble finding some that can be reproduced then maybe draw them yourselves. I think it's great that you've provided a glossary of terms, that's very useful but again I think there must be a formatting issue as there are strange boxes under the table. Also add an introduction to your page, to cover a basic overview of what will be contained within the project page and a summary of intergrins. In summary I think you've done some good work on the page, but some content still needs to have more info and detail. Once that is complete the page will be really great. Keep up the good work and good luck :).
I think this one of the the better pages that I've seen so far from all the others. The structure is laid out really well and the content is clear, informative and easy to understand. I think the amount of content is consistent and balanced for each sub topic. This is great because it's not too overwhelming to read and it's also interesting. Your referring is also done really well. I did notice some in text citations weren't in the correct form, I'd suggest changing them. Your images are really great, maybe try to add some more throughout the page especially in the clinical significance sub topic, to make it a bit more vibrant. Great work in having that drawing under the assembly of elastic fibers, looks good. Another suggestion is to maybe have a bit more info in the introduction as it seems a little small, but it's understandable that you haven't filled it out completely as its the part that you do at the very end. So once you've finished everything else maybe just add another paragraph to it to highlight what the page will cover and a general overview of the topic and your page. Overall the page has been set out really well there's nothing that really needs changing, so I don't have many suggestions to make. I think you should just add a few more image, drawings or tables here and your good to go.
Really great effort everyone ! :)
The layout of you page is effective and all the content provided is easy to understand. I’d consider having some more information in some sub topics especially abnormalities and current research. I am assuming you have the article references there because you’re going to still add information there. You’ve done great work and a lot of research is obviously seen from your extensive reference list. Your referencing is done really well, the in text citations and then just one reference list at the end is good. In your reference list there are empty spaces, just double check the formatting for those. Ensure that all your images do have a full reference included refer to the editing basics tab on the left hand side of the cell biology home page and scroll down into the images section. I’d suggest more images for all the sections, maybe a table for your history section. The images, tables and drawings will make the page more vibrant. Therefore I think your page is great so far, it just needs some more visuals to make it more colorful.
Great effort so far, keep up the good work :).
I think you’ve covered an extensive amount of information in the content you’ve provided. It is very obvious that you’ve all contributed equally and exceptionally well, based on the amount of info and research.The layout of your topics and subtopic looks good it’s clear and concise. I only have a few minor suggestions and these include having only one reference list at the end of the page. I think that maybe your unsure of how to do that so here are some tips. Under your reference heading type in < references/ > (without the spaces) and all the references used will be shown under that one heading. It will add more flow to the page. Any additional referencing help can be found in the editing basics tab and then click on project referencing. I’d also suggest making your images slightly smaller because they are quite large at the moment and they sort of take over the content. Also your Copyright information should be only in the image, don’t need to have it on the page too. Another suggestion is to maybe consider some drawings, tables or collapsable tables their great for condensing info. Otherwise I think your page is pretty much set, just adjust some minor formatting things here and there and your good. I think you’ve done a brilliant job, great work everyone !! ☺
You’ve structured the page well under the topics and sub topics that you currently have. It seems that there still need to be more content added onto some of the sub topics, especially at the beginning. I am sure you’ll have time to complete it all after these reviews. The information is researched well so far, its easy to understand. Consider having more images, maybe showing the formation of the collagen. Also some images under the abnormalities would be good too. If you have trouble finding images that can be reproduced, then draw them yourselves. Make sure you look over the referencing of all the images that you have so far, you don't want to loose marks for having some things missing. A formatting suggestion would be to have the antibodies under the ‘current research’ topic in a table or easy to read format. The group clearly knows how to cite all their content, the in text citations are all formatted really well and according to the correct formulas.The a reference list is correctly added. There are some references that have not been cited but I am assuming that this is because you are still going to add more content from the references. Make sure you add these to the reference list later on. The drawing of the articular cartilage is really good, clear and easy to understand. Overall the group has done some great work content wise and the layout of the page. I’d just suggest some more images, maybe tables and just fill in some of the missing content. Good work so far ☺.
During the lab
Human cell line
Medium: Mesenchymal Stem Cell Growth Kit
Mouse cell line
Medium: Dulbecco's modified Eagle's medium Dulbecco's modified Eagle's medium
1. Identify an antibody that can been used in your group's extracellular matrix project.
Antibody for Fibrillin the product is called Anti-Fibrillin 1 antibody [11C1.3]
2.Identify the species deriving the antibody.
3. Identify the working concentration for the antibody.
500 µl at 0.2 mg/ml
4.Identify a secondary antibody that could be used with this antibody.
Goat Anti-Mouse IgG H&L (DyLight® 488) preadsorbed (ab96879)
5.Identify a papers that has used this antibody.
<pubmed>20850568</pubmed> <pubmed>24190885</pubmed> <pubmed>19358237</pubmed>
Group 3 - Cytoskeleton exercise
1. Do you observe a change in morphology of differentiated B35 cells over-expressing Tm4 compared to undifferentiated B35 cells that do not over-express Tm4. If you see a change in morphology briefly describe the observed morphological change.
Yes, The control group; wild type undifferentiated B35 cells that were mostly seen include the 'broken fan' and 'stumped' phenotype (supported by the graph above). These consisted of very short lamellum lamelle and neurites. Although most of the TM4 over-expressed undifferentiated B35 cells were of the 'pronged' and 'stringed' phenotype. They had had multiple and longer neurites and single lamellum. There were no 'fan' or 'pygnotic' cell phenotypes observed, they were difficult to distinguish.
2. Provide a rationale that explains your observation on the molecular level.
- more actin in neurites - actin holds the lamellum lamelle, tropomyosin sits with the actin, it's intertwined - so the Tm4 helps to add actin which then is responsible for the 'pronged' and 'stringed' phenotypes, it allows for more stability to occur
<pubmed>15086897</pubmed> The paper discusses Autosomal dominant Alport syndrome and it’s relation to the COL4A3 or COL4A4 gene. The syndrome is identified as a heterogenous nephropathy both clinically and genetically, it is also associated with ocular lesions or deafness . Mutations of COL4A5 genes are transmitted as X-linked semi-dominant , and so males are more likely to be affected. The method involved assessing the autosomal dominant Alport syndrome among several families that were predisposed to the gene. The technique used to identify the COL4A3 and COL4A4 genes was high performance liquid chromatography (HPLC). The mutations was then evaluated using automated sequencing. All members had their blood samples collected and peripheral blood leukocytes were used to isolate genomic DNA. From the results it was concluded that two of the families expressed the COL4A4 gene and the other COL4A3 gene. The overall assessment highlighted that autosomal dominant Alport syndrome patients have a wide range of clinical presentations among these include end staged renal disease, nonprogressive isolated microhematuria or no symptoms at all.
Thin Glomerular Basement Membrane
<pubmed>16467446</pubmed> The common cause of hematuria is thin basement membrane nephropathy, other causes may include Alport syndrome and IgA nephropathy. Through electron microscopy it has been noted that patents with thin basement membrane nephropathy have normal renal function, negligible proteinuria and their glomerular basement membranes are uniformly thinned. The disease is believed to be linked to a IV collagen trimer. Autosomal thin basement membrane nephropathy has evidently been caused by COL4A3 or COL4A4 heterozygous mutations. However combined heterozygous or homozygous mutations show autosomal recessive Alport syndrome being expressed in the same genes. The paper outlines key characteristics of the nature of the disorder including epidemiology, clinical features, renal biopsy, genetics, pathogenesis, diagnosis and finally management. It provides an insightful and in depth understanding of the abnormality.
<pubmed>PMC1892429</pubmed> The absence or incomplete basement membrane of blood vessels has been found in many tumours. In tumors blood vessels often have several function and structural abnormalities, they consist of mural cells (composed of smooth muscle cells or pericytes), endothelial and the basement membrane. This study focuses on the abnormalities of blood vessel basement membrane and endothelial sprouts in models of mouse tumors. Immunohistochemistry was used to localise markers of endothelial cells and basement membrane in order to fins out the allocation and extent of coverage of vascular basement membrane. Then the linkages between pericytes, endothelial cells and the vascular basement membrane were observed. The final objective was to determine if endothelial sprouts were related to basement membrane. Three tumor mouse models were examined these included; Lewis lung carcinomas, spontaneous pancreatic islet and mammary carcinomas .The method involved immunoflurescence histochemistry, transmission EM and morphometric measurements. From the results it was concluded that in tumors the vascular basement membrane shows significant abnormalities these included multiple layers, loose association with pericytes and endothelial cells, and irregular thickness. The endothelial sprouts were covered by the basement membrane, such features have proved to be consistent with pericytes and endothelial cells among tumors.
Dystrophic Epidermolysis Bullosa
<pubmed>PMC4319308</pubmed> Dystrophic Epidermolysis Bullosa (RDEB) consists of several types including the recessive; where mutations are present in both alleles of the type VII collagen gene. The disease involves blistering of the skin which is intractable genetically. In the disease there is loss of genetic function in functional type VII collagen in the cutaneous basement zone. The outcome that is presented is a separation of epithelial structures from underlying dermis. To combat this many studies have focused on allogenic bone marrow transplantations (BMT) which can improve the skin blistering phenotype of RDEB among patients by restoring the functional type VII collagen. This study used mice models to present their investigation. The methods included bone marrow transplants, skin graft models, ELISA , Immunofluorescent microscopy, flow cytometry & cell sorting, PCR, CXCR4 antagonist delivery and statistical data. Furthermore from the results the study was able to show that transplanted bone marrow-derived PDGFRa+ mesenchymal cells are able to supplement functional type IV collagen in the basement membrane zone as the cells migrate to the RDEB mouse skin. So bone marrow derived mesenchymal cells could be a supposed source of functional type IV collagen in RDEB patient skin. Although these finds would need to be further investigated in a human setting, to be considered for clinical applications.
During the lab
Search term: membrane Basement membrane
<pubmed limit=5>Basement membrane</pubmed>
Identify a research article (not review) on super resolution microscopy and prepare a brief (1-2 paragraph) summary of the paper. Add the referenced summary to your page.
This research paper focuses on natural killer cell synapses on the supported lipid bilayer. The use of glass supported planar lipid bilayer (SLB) has been successful in the immunological synapse formation. This is as a result of the formation of the horizontal interface due to the glass supported planar lipid bilayer’s ability to emulate a target cell surface. The development in super resolution imaging has provided an innovation to more specific details of the synapse structure. The structure of natural killer cell synapses on supported lipid bilayers can be observed with the aid of a new technique known as stimulated emission depletion (STED). This technique is among many used in conjunction to super resolution imagery. It has become one of the newest and most sought after innovations in microscopy imaging all over the world.
In this case the technique has been beneficial in providing more detail for immunological synapses. The study uses the super resolution imaging with the STED technique utalising a depletion laser. This new laser is able to show an extensive collection of fluorescent dyes. The method involved preparation of liposomes, dialysis of liposomes, determining antibody density of lipid bilayers, isolating and culturing human NK cells. Then assembling glass-supported planar lipid bilayer’s and finally the imaging of NK synapse on lipid bilayer. The paper also provides a video to support the method of the procedures involved in the techniques.
The protocol involved a large amount of adaptations and the study involved SLB techniques and STED which have proven to show basic lipid dynamics, tumor cell metastasis, formation of synapses and intracellular signaling. The results provided a preparation for liposomes, determining protein density on lipid bilayers, construction of protein embedded bilayers and super resolution images using STED. This will prove to be an incredible insight into a wide range of future studies among many fields.
During the lab
Mycobacterium smegmatis LS2 strain
- ---- - add website here - ---
Mycobacterium smegmatis LS2 strain
Electron micrographs of Myobacterium smegmatis LS2 strain during shifting temperature changes from 30°C to 42°C. Figures A, B & C showed growth at 30°C from the size and shape of Myobacterium smegmatis LS2 strain. They were similar to the wild type strain.
Figures D, E & F showed that at 42°C Myobacterium smegmatis LS2 strains had a rougher cell surface, lysis and the shapes were longer with bulb heads.
Figures H & I had a similar appearance to the previous figures at 42°C.
Figure G at 42 °C this wild type Myobacterium smegmatis had small oval shapes.
<pubmed>PMC3414508</pubmed>| PLoS One.
Shi W, Xu B, Wu L-M, Jin R-T, Luan H-B, et al. (2014) Oocytes with a Dark Zona Pellucida Demonstrate Lower Fertilization, Implantation and Clinical Pregnancy Rates in IVF/ICSI Cycles. PLoS ONE 9(2): e89409. doi:10.1371/journal.pone.0089409
Li S, Kang J, Yu W, Zhou Y, Zhang W, et al. (2012) Identification of M. tuberculosis Rv3441c and M. smegmatis MSMEG_1556 and Essentiality of M. smegmatis MSMEG_1556. PLoS ONE 7(8): e42769. doi:10.1371/journal.pone.0042769
© Li et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
- Note - This image was originally uploaded as part of a student project and may contain inaccuracies in either description or acknowledgements. Please contact the site coordinator if the uploaded content does not meet the original copyright permission or requirements, for immediate removal.