2 D-Electrophoresis Separation of the Wheat Leaf Proteome – Lab Report Example
2 D-electrophoresis separation of the wheat leaf proteome College: Results In the experiment, the aim was to carry out a 2 D-electrophoresis separation of the wheat leaf proteome. There are two gels produced in the experiment, in figure 1 the gel was ideal as it is evident by the spots that represents the proteins. In figure 2, the gel was not produced as expected; it was black with no spots. This means the gel was bad and it is recommended that the experiment to be run again.
Figure 1.2D gel that was considered to be ideal.
Figure 2.2D gel that was considered not good or ideal.
By definition, proteome is the entire protein complement that includes the modifications made to a give set of proteins by a cellular system or an organism (Ferry et al. 2002). Basing on the results of figure 1(2 D ideal gel), it is observed that the proteins were separated as expected; this implies that all the conditions were carefully maintained and the laboratory procedures were also followed accordingly. The dots indicate that the proteins were separated and their kDa could be identified from the left side using a ladder or the marker. In this case, the protein of interest could be identified, and then cut out from the strip so that it could be measured by spectrophotometer. At this point, the OD is then determined which could be extrapolated on the standard curve to get the concentration of this protein of interest. The concentration could be got by using the equation of the standard curve, hence calculating X value. The dark staining in this ideal gel could be due incomplete solubilization of the sample before application.
In figure 2,it is observed that there were no dots in the gel (no proteins on the gel) indicating that the experiment may have not been carried out accordingly.This could be due to the conditions of the protocal were not adhered to.It is known that the same sample used to produce results as shown in figure 1 was the one used to produce the results in figure 2.In this regard, the black gel could be due to the samples did not completely solubilized before application.In this case,it is necessary to ensure that the samples are stably and completely solubilized.Repeated precipitation-resolubilization cycles results to horizontal streaking.The samples could be poorly soluble in the rehydration solution:In this case the solubilizing components concentration shoul d be increased. Also increase the IPG buffer cncentration.In order to produce expected results it is recommended to trouble shout and identify why the results came out as what was not expected then follow the right procedure to generate the expected results
Proteomics is known to be a large scale study of protein functions and characteristics. The aim of proteomics research involves obtaining an integrated view of abnormal and normal organism or cellular processes at their constituent proteins level, for instance, on the basis of post translational modifications, protein-protein interactions, protein abundance as well as their regulatory networks (Ferry et al. 2002). Proteomics is classified into 2 major subgroups: Protein profiling that includes identification of specific markers and targets as well as functional proteomics that include the structure definition, function and interactions. 2-D PAGE is mostly suitable for studies related to protein profiling. It is also suitable to targeted or functional proteomic research, where the modification or expression of the given proteins is followed in the course of the alteration in growth conditions or systematic treatment regimens (Ferry et al. 2002)
Protein profiling such as the one carried out in the lab entails 2-D gels comparison in order to understand different biological processes through the determination of the absence or presence, modification states and down or up regulation of proteins. In this case, Profiling, hence acts as a technique of ‘‘discovering putatively causal correlations’’ between modification states or protein abundance and biological processes that are of interest. The aim of functional proteomics is to test predictions that are specific through a detailed proteins structures analysis, cellular locations, roles as well as interactions with other proteins. In this experiment, the aim was to determine if the wheat plant was resistant to the aphids through protein profiling. In this case, 2D electrophoresis was employed.
Ferry N, Stavroulakis S, Guan WZ, Davison GM, Bell HA, Weaver RJ, Down RE, Gatehouse JA, Gatehouse AMR.(2002).Molecular interactions between wheat and cereal aphid (Sitobion avenae): Analysis of changes to the wheat proteome. Proteomics 2011, 11(10).