User:Z5011413

From CellBiology

Lab 1

For today's attendance. --Z5011413 (talk) 15:47, 13 March 2014 (EST)

Lab 2

--Z5011413 (talk) 15:07, 20 March 2014 (EST)

Sub-heading

PubMed

ruff ruff

this is so great

Reference

This is prokaryotes.[1]

Lars Witting The natural selection of metabolism and mass selects lifeforms from viruses to multicellular animals. Ecol Evol: 2017, 7(21);9098-9118 PubMed 29152201

Conte Antonella, Papale Maria, Amalfitano Stefano, Mikkonen Anu, Rizzo Carmen, De Domenico Emilio, Michaud Luigi, Lo Giudice Angelina Bacterial community structure along the subtidal sandy sediment belt of a high Arctic fjord (Kongsfjorden, Svalbard Islands). Sci. Total Environ.: 2017, 619-620;203-211 PubMed 29149744

Messaouda Khallef, Süleyman Cenkci, Dilek Akyil, Arzu Özkara, Muhsin Konuk, Djamel Eddine Benouareth Ames and random amplified polymorphic DNA tests for the validation of the mutagenic and/or genotoxic potential of the drinking water disinfection by-products chloroform and bromoform. J Environ Sci Health A Tox Hazard Subst Environ Eng: 2017;1-6 PubMed 29148923

Ulrich Omasits, Adithi R Varadarajan, Michael Schmid, Sandra Goetze, Damianos Melidis, Marc Bourqui, Olga Nikolayeva, Maxime Québatte, Andrea Patrignani, Christoph Dehio, Juerg E Frey, Mark D Robinson, Bernd Wollscheid, Christian H Ahrens An integrative strategy to identify the entire protein coding potential of prokaryotic genomes by proteogenomics. Genome Res.: 2017; PubMed 29141959

Esti Singer, Yardena Bh Silas, Sigal Ben-Yehuda, Ophry Pines Bacterial fumarase and L-malic acid are evolutionary ancient components of the DNA damage response. Elife: 2017, 6; PubMed 29140245


  1. Hao Tong, Jan Mrázek Investigating the interplay between nucleoid-associated proteins, DNA curvature, and CRISPR elements using comparative genomics. PLoS ONE: 2014, 9(3);e90940 PubMed 24595272


The Golgi Apparatus

AFM images of Golgi apparatus.

Golgi Apparatus Images.png

Reference

Haijiao Xu, Weiheng Su, Mingjun Cai, Junguang Jiang, Xianlu Zeng, Hongda Wang The asymmetrical structure of Golgi apparatus membranes revealed by in situ atomic force microscope. PLoS ONE: 2013, 8(4);e61596 PubMed 23613878

| PLoS One.

Copyright

© 2013 Xu 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.

Citation: Xu H, Su W, Cai M, Jiang J, Zeng X, et al. (2013) The Asymmetrical Structure of Golgi Apparatus Membranes Revealed by In situ Atomic Force Microscope. PLoS ONE 8(4): e61596. doi:10.1371/journal.pone.0061596


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.


The Mammalian Interphase Nucleus

The mammalian interphase nucelus.png

The diversity in size and number of some of the major functional nuclear compartments is shown in the image.


Reference

Carol Shiels, Niall M Adams, Suhail A Islam, David A Stephens, Paul S Freemont Quantitative analysis of cell nucleus organisation. PLoS Comput. Biol.: 2007, 3(7);e138 PubMed 17676980

| PLoS One. Journal.


Copyright

© 2007 Shiels 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.

Citation: Shiels C, Adams NM, Islam SA, Stephens DA, Freemont PS (2007) Quantitative Analysis of Cell Nucleus Organisation. PLoS Comput Biol 3(7): e138. doi:10.1371/journal.pcbi.0030138

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.
The {{The Mammalian Interphase Nucleus.}}

3D Multicolor Super-Resolution Imagine Offers Improved Accuracy in Neuron Tracing

Melike Lakadamyali, Hazen Babcock, Mark Bates, Xiaowei Zhuang, and Jeff Lichtman in their experiment used a type of super-resolution microscopy called stochastic optical reconstruction microscopy (STORM), for tracing neural connectivity using cultured hippocampal neurons obtained from wild-type neonatal rat embryos as a model system. With the advanced 3D resolution, they could distinguish and trace substantially more neuronal process in the super-resolution images rather than confocal images. The accuracy was even further improved by using a multicolor version.

Reference

Melike Lakadamyali, Hazen Babcock, Mark Bates, Xiaowei Zhuang, Jeff Lichtman 3D multicolor super-resolution imaging offers improved accuracy in neuron tracing. PLoS ONE: 2012, 7(1);e30826 PubMed 22292051

| [1]


Copyright

©2012 Lakadamyali 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.

Citation: Lakadamyali M, Babcock H, Bates M, Zhuang X, Lichtman J (2012) 3D Multicolor Super-Resolution Imaging Offers Improved Accuracy in Neuron Tracing. PLoS ONE 7(1): e30826. doi:10.1371/journal.pone.0030826

Lab 3

--Z5011413 (talk) 16:30, 27 March 2014 (EST)

Article Reviews for Project

  • Materials for potential import into the mitochondria are recognized by a specific receptor on the surface of the mitochondria that delivers them into a protein import channel in the outer membrane. The receptor and channel include several non-identical subunits and are referred to as the Tom Complexes (“Transport across the Outer Membrane”). The receptor consists of at least four non-identical integral membrane proteins named (after their approximate molecular masses in kDa) Tom70, Tom37, Tom22, and Tom20. This articles goes through the functions of these proteins but only Tom20 is essential for protein transport. [1]
  • The two receptors, Tom22 and Tom20, are critical for the assembly of Tom40 channel units and have distinct roles as organizers of Tom40 dimers into larger TOM structures. Based on biochemical and electron microscopy studies of the TOM complex isolated from yeast mitochondria, this article discovered the molecular reason for the different number of channel-like structures. The TOM complex from wild-type yeast contains up to three stain-filled centers, while from a mutant yeast selectively lacking Tom20, the TOM complex particles contain only two channel-like structures. From mutant mitochondria lacking Tom22, native electrophoresis separates an ∼80 kDa subcomplex that consists of Tom40 only and is functional for accumulation of a precursor protein. The article concludes that while Tom40 forms the import channels, the two receptors Tom22 and Tom20 are required for the organization of Tom40 dimers into larger TOM structures. [2]
  • In this article Rapaport obtained knowledge about the TOM complex through studying Saccharomyces cerevisiae and Neurospora crassa that contain homologous components to higher organisms like mammals and plants. The receptors Tom20 and Tom22 are involved in the translocation of most protein precursors, especially those with N-terminal targeting signals. Tom70, forms a binding site for a more bound set of molecules that will become proteins, most notably the mitochondrial carrier family. The subunits Tom40, Tom22, Tom7, Tom6 and Tom5 are embedded in the outer membrane where they form the Tom core complex, also called "the general insertion pore". Tom40 forms the protein-conducting channel, whereas Tom22 and Tom5 probably link the receptors to the pore (this is not concluded yet). In conclusion, the article found that Tom40 is essential for the viability of yeast and N. crassa cells. Although Tom22 was also thought to be essential, it has recently been reported that, under special genetic conditions, a very ‘sick’ tom22Δ strain can be isolated. None of the other Tom subunits is essential for viability in yeast or N. crass. [3]
  • In the article, “Biogenesis of Porin of the Outer Mitochondrial Membrane Involves an Import Pathway via Receptors and the General Import Pore of the Tom Complex”, the mitochondira from Neurospora crassa and Saccharomyces cerevisiae were used to analyze the import pathway of porin (voltage dependent ion channel) which is the most abundant protein of the mitochondrial outer membrane. The process of import and assembly of the protein is known to be dependent on the surface receptor Tom20, but the requirement for other mitochondrial proteins were explored in this article. [4]

Image Upload on Tom Complex

The Translocase of the outer mitochondria membrane(TOM)

The TOM complex contains the initial receptors Tom20 and Tom70 (yellow). These two receptors are loosely attached to the rest of the complex (the TOM core complex); using mild detergent, they purify with the other Tom components, but not under more rigorous conditions. The TOM core complex builds the protein-conducting pore and contains Tom5, Tom6, Tom7, Tom22 and Tom40. The names of the Tom subunits reflect their molecular weights. [5]

  1. G Schatz The protein import system of mitochondria. J. Biol. Chem.: 1996, 271(50);31763-6 PubMed 8943210
  2. Kirstin Model, Thorsten Prinz, Teresa Ruiz, Michael Radermacher, Thomas Krimmer, Werner Kühlbrandt, Nikolaus Pfanner, Chris Meisinger Protein translocase of the outer mitochondrial membrane: role of import receptors in the structural organization of the TOM complex. J. Mol. Biol.: 2002, 316(3);657-66 PubMed 11866524
  3. Kai Stefan Dimmer, Doron Rapaport The enigmatic role of Mim1 in mitochondrial biogenesis. Eur. J. Cell Biol.: 2009, 89(2-3);212-5 PubMed 19944477
  4. T Krimmer, D Rapaport, M T Ryan, C Meisinger, C K Kassenbrock, E Blachly-Dyson, M Forte, M G Douglas, W Neupert, F E Nargang, N Pfanner Biogenesis of porin of the outer mitochondrial membrane involves an import pathway via receptors and the general import pore of the TOM complex. J. Cell Biol.: 2001, 152(2);289-300 PubMed 11266446
  5. Kai Stefan Dimmer, Doron Rapaport The enigmatic role of Mim1 in mitochondrial biogenesis. Eur. J. Cell Biol.: 2009, 89(2-3);212-5 PubMed 19944477

Lab 4

--Z5011413 (talk) 15:11, 3 April 2014 (EST)

Anitbody against Cadherin: N-Cadherin Monoclonal Antibody

(class exercise)

  • type of antibody- monoclonal
  • species raised in- mouse
  • species reacts against- chicken, monkey, rabbit, human, mouse, rat
  • types of application uses- electron microscopy, flow cytometry, immunocytochemistry, immunohistochemistry, and western blot
  • reference using N-Cadherin antibody- immunohistochemistry: 10-20 μg/mL using rat cardiac muscle frozen sections and western blot: 10-20 μg/mL using chicken cardiac muscle or COS-7 cell extracts
  • price- 411.60 (AUD)

Reference: Sigma-Aldrich

Anti-Tim23 Antibody

  1. TIM23 Antibody (T-20)
  2. T-20 is a goat polyclonal IgG
  3. The working concentration for this antibody is 200 µg/ml
  4. Secondary antibody for IgG would be ABC-elite reagent
  5. This paper used the T-20 Antibody: “Tim23–Tim50 pair coordinates functions of translocators and motor proteins in mitochondrial protein import” [1]


Reference: [2]

  1. Yasushi Tamura, Yoshihiro Harada, Takuya Shiota, Koji Yamano, Kazuaki Watanabe, Mihoko Yokota, Hayashi Yamamoto, Hiromi Sesaki, Toshiya Endo Tim23-Tim50 pair coordinates functions of translocators and motor proteins in mitochondrial protein import. J. Cell Biol.: 2009, 184(1);129-41 PubMed 19139266


Lab 5

--Z5011413 (talk) 15:16, 10 April 2014 (EST)

Lab6

attendance --Z5011413 (talk) 15:08, 17 April 2014 (EST)

Exercise 1 Graph

Undifferentiated B35 Cell Count.PNG This above experiment was specifically dealing with Tropomyosin 4.

Lab7

attendance --Z5011413 (talk) 16:40, 1 May 2014 (EST)

Lab 9

attendance --Z5011413 (talk) 15:12, 15 May 2014 (EST)

Apoptosis Exercise

Hypothesis: Isolated cells grown in optimal tissue culture conditions will most likely undergo apoptosis once placed in a foreign environment.

Aim: To determine whether the cell undergoes an intrinsic or extrinsic apoptotic pathway by observing the amount of caspase 8 activated.

Method: Human adherent cells are used for the tissue culture. They are washed carefully and loose cells are isolated. Adherent cells are gently spun and labelled with FAM green fluorescence from the FLICATM kit before trypsinization. Cells are incubated with the FLICA at 37oC for roughly 60 minutes before washing. After the addition of apoptosis wash buffer, cells are placed on ice and protected from light. Observations are made by the analysis of cells with a flow cytometer. Click ‘Protocol’ tab in the following link (Immunochemistry Technologies) for more detailed instructions.

Suppliers: Immunochemistry Technologies [3]Resource: Green FLICA™ Caspase 8 Assay Kit

Student Made Flow Chart: Apoptosis flow diagram.PNG

Results:If there is a high fluorescence reading observed, that means that there is a high amount of caspase 8 present and therefore, leads to an extrinsic apoptotic pathway. However, if there is a low fluorescence reading observed, there is less caspase 8 and hence, there is no extrinsic pathway activated.

Peer Review for Group Projects

Group 1

-You all are focusing on specifically phagocytosis and your topic is endocytosis. I have no idea if you have talked this over with Mark but perhaps you should briefly talk about endocytosis and the other mechanisms it includes besides phagocytosis?

-could benefit from the addition of a glossary at the end clearly defining scission, lysosome, phaolysosome, PMNs, etc.

-headings/sections could be set up in a better format

  • ‘Diseases’ and ‘Current/future Research’ sections should be integrated with the rest of the content
  • Maybe the overall mechanism of phagocytosis should be put before the more specific Receptors section

-The description under the ‘Receptors’ section gives a great overall discussion in the realm of phagocytosis. This paves the way for the sub-sections below it on specific receptor-mediated phagocytosis. This is a great idea and maybe you should add an overall discussion for your ‘Mechanism of Phagocytosis’ as well.

-phagocytosis is a really interesting action to see at the microscopic level so perhaps a video showing an antigen being eaten up would be cool! I think Mark showed us one during class if you are allowed to use that?

-some content seems to be referenced correctly but make sure all references are at the bottom and not dispersed throughout

-This project has a great foundation of information- the overall meat of phagocytosis is there but the organization of the content needs to be adjusted.

Group 3

-more content should be added to the blank sections obviously; when you write your intro make sure to briefly touch on the major components of your project like RanGTP/GDP, IMP and Ran-independent nuc. Import pathways, diseases, etc.

-the content cited looks very professional and clean; great job in referencing the pubmed ID for each, great sources

-why are there pictures in your references section? Is this a glitch or are you further referencing these?

-a student drawing on the basic structure on a nucleoporin would aid the discussion greatly, actually pictures of the structures listed in your project would be helpful in understanding their function

-abbreviations are a little unclear, maybe the addition of a glossary with discriptions of these would be beneficial

-I like how you used numbers to clearly point out the four different NPC Models

-the overall format of this project flows in a way that makes it easy to understand but maybe introductions to each sub-topic in the project would make it even better, for example an introduction on the nuclear pore complex before jumping into NPC Models

Group 4

-the addition of images would make the processes you are describing much more clear

-the content lacks transitional phrases and thus reads very “choppy”; try to make it flow and all connect

-the points relating to transport from neuron soma to processes are not well defined, instead of defining everything about kinesin focus more on the actual processes of transport

-a clear direction you are heading with the use of organized headings and subheadings should be added

-when you all were presenting it was pointed out that your content was moving in the wrong direction so it is difficult to give details about the body of your project but you still have a lot of time to make it work ☺ best of luck!

Lab 10

attendance--Z5011413 (talk) 15:20, 22 May 2014 (EST)