Showing papers on "Gel electrophoresis published in 2006"
TL;DR: This protocol is for the in-gel digestion of both silver and Coomassie-stained protein spots or bands and can be followed by MALDI-MS or LC-MS/MS analysis to identify proteins at sensitivities better than a few femtomoles of protein starting material.
Abstract: In-gel digestion of proteins isolated by gel electrophoresis is a cornerstone of mass spectrometry (MS)-driven proteomics. The 10-year-old recipe by Shevchenko et al. has been optimized to increase the speed and sensitivity of analysis. The protocol is for the in-gel digestion of both silver and Coomassie-stained protein spots or bands and can be followed by MALDI-MS or LC-MS/MS analysis to identify proteins at sensitivities better than a few femtomoles of protein starting material.
4,493 citations
TL;DR: Cross‐linked homogeneous 5 per cent polyacrylamide gel in rod form is a convenient medium for sharp analytical electrophoretic separation in microgram quantities of most serum proteins and of many biological ampholytes.
Abstract: Summary
Cross-linked homogeneous 5 per cent polyacrylamide gel in rod form is a convenient medium for sharp analytical electrophoretic separation in microgram quantities of most serum proteins and of many biological ampholytes.
532 citations
TL;DR: FA-nanoparticles have lower cytoxicity, good DNA condensation, positive zeta potential and particle size around 118 nm, which makes them a promising candidate as a non-viral gene vector.
373 citations
TL;DR: It is, however, beyond the scope of this paper to consider the theoretical problems of electrophoresis as modified by the filtration effect, and this paper is limited to the applications of gel electrophoreis and to the problems the authors have encountered with it since its first publication in 1959.
Abstract: This symposium on gel electrophoresis includes in its program papers on starch gel electrophoresis, agar gel electrophoresis, acrylamide gel electrophoresis, and the specialized technique of disc electrophoresis. It is appropriate to consider what characteristic distinguishes gel electrophoresis from other techniques and makes it especially useful in the analysis of protein and other complex mixtures. In my opinion it is the “sieve” or “molecular filtration” effect, first described by 0. Smithies,’ that is the essential characteristic of gel electrophoresis. The combination of electrophoretic effects and molecular filtration effects produces new and startling resolutions that cannot be obtained by other electrophoretic procedures. This combination‘of effects is not present in agar gel electrophoresis. Therefore agar techniques should logically be classified with paper electrophoresis techniques, which they most closely resemble with respect to the type of pattern produced. The specialized technique of “disc electrophoresis” intrinsically applies a discontinuous buffer system to sharpen the electrophoretic zones of the individual components of a mixture but does not in itself make use of the special feature of gel electrophoresis, which is the molecular filtration effect. Immunoelectrophoresis is also a specialized technique, namely a technique of detecting and identifying the zones separated by electrophoresis. It could equally well be applied to a paper electrophoresis type of separation, as it is in agar gel, or to a molecular filtration type of separation as is produced by starch gel or acrylamide gel. These considerations provide a rational basis for focusing our attention on the special problems that have arisen in gel electrophoresis (defined as electrophoresis technique combining both electrophoretic and molecular filtration effects) and the special results available from it. It is, however, beyond the scope of this paper to consider the theoretical problems of electrophoresis as modified by the filtration effect. Instead, this paper is limited to the applications of gel electrophoresis and to the problems we have encountered with it since our first publication in 1959.2 It is evident from a quick perusal of the program that we are devoting most of our attention to the disc electrophoresis technique of Ornstein. Since, apparently, I am the only speaker to describe the vertical slab technique I must begin with a description of our procedure and apparatus. Our procedure starts with a vertical gel slab prepared in a specially designed cell (FIGURE combining gel mold, buffer reservoirs and electrodes, and cooling plates all in one unit. We prefer a flat slab in contrast to a cylindrical gel for several reasons. First, the flat slab provides maximum surface area for cooling the gel. Second, the resulting patterns are easier to quantitate in standard recording densitometers. Third, a large number of samples can be processed in a single gel, making the technique easier to carry out in the laboratory and facilitating the direct intercomparison of specimens processed under identical conditions all the way through to the final densitometer recording. Fourth, and most important, the flat slab permits the application of two-dimensional techniques, which is impossible in the vertical tube apparatus. We have never observed any difficulties with stress relief or separation from
270 citations
TL;DR: MudPIT was used to analyse the ‘tightly‐bound’ proteome for purified cell walls from Arabidospis cell suspension cultures and revealed that a large proportion of the proteins were enzymes, notably carbohydrate active enzymes, peroxidases and proteases.
Abstract: With the completion of the sequencing of the Arabidopsis genome and the recent advances in proteomic technology, the identification of proteins from highly complex mixtures is now possible. Rather than using gel electrophoresis and peptide mass fingerprinting, we have used multidimensional protein identification technology (MudPIT) to analyse the "tightly-bound" proteome for purified cell walls from Arabidopsis cell suspension cultures. Using bioinformatics for the prediction of signal peptides for targeting to the secretory pathway and for the absence of ER retention signal, 89 proteins were selected as potential extracellular proteins. Only 33% of these were identified in previous proteomic analyses of Arabidopsis cell walls. A functional classification revealed that a large proportion of the proteins were enzymes, notably carbohydrate active enzymes, peroxidases and proteases. Comparison of all the published proteomic analyses for the Arabidopsis cell wall identified 268 non-redundant genes encoding wall proteins. Sixty of these (22%) were derived from our analysis of tightly-bound wall proteins.
239 citations
TL;DR: This chapter describes how a combination of high-performance liquid chromatography (HPLC) and exoglycosidase enzyme array digestions can deliver quantitative glycan analysis of sugars released from glycoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel bands by matching HPLC elution positions with a database of standard glycans.
Abstract: In contrast to the linear sequences of protein and DNA, oligosaccharides are branched structures. In addition, almost all glycoproteins consist of a heterogeneous collection of differently glycosylated variants. Glycan analysis therefore requires high-resolution separation techniques that can provide detailed structural analysis, including both monosaccharide sequence and linkage information. This chapter describes how a combination of high-performance liquid chromatography (HPLC) and exoglycosidase enzyme array digestions can deliver quantitative glycan analysis of sugars released from glycoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel bands by matching HPLC elution positions with a database of standard glycans.
233 citations
TL;DR: In this paper, the effect of storage time and temperature on the solubility of milk protein concentrate (MPC85) was investigated using solubilities tests, gel electrophoresis and mass spectrometry.
217 citations
TL;DR: DIGE combined with digital image analysis greatly improves the statistical assessment of proteome variation and is capable of reliably detecting as little as 0.5 fmol of protein, and protein differences down to ± 15%, over a >10,000-fold protein concentration range.
Abstract: Two-dimensional difference gel electrophoresis (2D DIGE) is a modified form of 2D electrophoresis (2DE) that allows one to compare two or three protein samples simultaneously on the same gel. The proteins in each sample are covalently tagged with different color fluorescent dyes that are designed to have no effect on the relative migration of proteins during electrophoresis. Proteins that are common to the samples appear as 'spots' with a fixed ratio of fluorescent signals, whereas proteins that differ between the samples have different fluorescence ratios. With the appropriate imaging system, DIGE is capable of reliably detecting as little as 0.5 fmol of protein, and protein differences down to +/- 15%, over a >10,000-fold protein concentration range. DIGE combined with digital image analysis therefore greatly improves the statistical assessment of proteome variation. Here we describe a protocol for conducting DIGE experiments, which takes 2-3 d to complete.
217 citations
TL;DR: Triton X-100 was used in combination with various concentrations of NaCl to separate intact WSSV virions into distinct fractions such that each fraction contained envelope and tegument proteins, Tegument and nucleocapsid proteins, or nucleocapid proteins only.
Abstract: The protein components of the white spot syndrome virus (WSSV) virion have been well established by proteomic methods, and at least 39 structural proteins are currently known. However, several details of the virus structure and assembly remain controversial, including the role of one of the major structural proteins, VP26. In this study, Triton X-100 was used in combination with various concentrations of NaCl to separate intact WSSV virions into distinct fractions such that each fraction contained envelope and tegument proteins, tegument and nucleocapsid proteins, or nucleocapsid proteins only. From the protein profiles and Western blotting results, VP26, VP36A, VP39A, and VP95 were all identified as tegument proteins distinct from the envelope proteins (VP19, VP28, VP31, VP36B, VP38A, VP51B, VP53A) and nucleocapsid proteins (VP664, VP51C, VP60B, VP15). We also found that VP15 dissociated from the nucleocapsid at high salt concentrations, even though DNA was still present. These results were confirmed by CsCl isopycnic centrifugation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry, by a trypsin sensitivity assay, and by an immunogold assay. Finally, we propose an assembly process for the WSSV virion.
203 citations
TL;DR: Altering in the root proteome of hydroponically grown Arabidopsis thaliana plants treated with 10 microM Cd(2+) for 24 h triggers the synthesis of phytochelatins, glutathione-derived metal-binding peptides, shown here as PC2 accumulation.
Abstract: Cadmium is a major environmental pollutant that enters human food via accumulation in crop plants. Responses of plants to cadmium exposure--which directly influence accumulation rates--are not well understood. In general, little is known about stress-elicited changes in plants at the proteome level. Alterations in the root proteome of hydroponically grown Arabidopsis thaliana plants treated with 10 microM Cd(2+) for 24 h are reported here. These conditions trigger the synthesis of phytochelatins (PCs), glutathione-derived metal-binding peptides, shown here as PC2 accumulation. Two-dimensional gel electrophoresis using different pH gradients in the first dimension detected on average approximately 1100 spots per gel type. Forty-one spots indicated significant changes in protein abundance upon Cd(2+) treatment. Seventeen proteins found in 25 spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Selected results were independently confirmed by western analysis and selective enrichment of a protein family (glutathione S-transferases) through affinity chromatography. Most of the identified proteins belong to four different classes: metabolic enzymes such as ATP sulphurylase, glycine hydroxymethyltransferase, and trehalose-6-phosphate phosphatase; glutathione S-transferases; latex allergen-like proteins; and unknown proteins. These results represent a basis for reverse genetics studies to better understand plant responses to toxic metal exposure and to the generation of internal sinks for reduced sulphur.
200 citations
Journal Article•
TL;DR: With fluorescently labeled primers and dideoxynucleotide DNA sequencing, it is shown that the terminal base of each digested fragment may be accurately identified with a capillary-based instrument.
Abstract: We have adapted the techniques of DNA footprint analysis to an Applied Biosystems 3730 DNA Analyzer. The use of fluorescently labeled primers eliminates the need for radioactively labeled nucleotides, as well as slab gel electrophoresis, and takes advantage of commonly available automated fluorescent capillary electrophoresis instruments. With fluorescently labeled primers and dideoxynucleotide DNA sequencing, we have shown that the terminal base of each digested fragment may be accurately identified with a capillary-based instrument. Polymerase chain reaction (PCR) was performed with a 6FAM-labeled primer to amplify a typical target promoter region. This PCR product was then incubated with a transcriptional activator protein, or bovine serum albumin as a control, and then partially digested with DNase I. A clone of the promoter was sequenced with the Thermo Sequenase Dye Primer Manual Cycle Sequencing kit (USB) and the FAM-labeled primer. Through the use of Genemapper software, the Thermo sequenase and DNasei digestion products were accurately aligned, providing a ready means to assign correct nucleotides to each peak from the DNA footprint. This method was used to characterize the binding of two different transcriptional activator proteins to their respective promoter regions.
TL;DR: The activity of the purified CNV replicase correlated with viral RNA replication in the above-mentioned ssa1 ssa2 mutant and in the Ssa overexpression strains, suggesting that Ssa1/2p likely plays an important role in the assembly of the CNV Replicase.
Abstract: Plus-strand RNA virus replication occurs via the assembly of viral replicase complexes involving multiple viral and host proteins. To identify host proteins present in the cucumber necrosis tombusvirus (CNV) replicase, we affinity purified functional viral replicase complexes from yeast. Mass spectrometry analysis of proteins resolved by two-dimensional gel electrophoresis revealed the presence of CNV p33 and p92 replicase proteins as well as four major host proteins in the CNV replicase. The host proteins included the Ssa1/2p molecular chaperones (yeast homologues of Hsp70 proteins), Tdh2/3p (glyceraldehyde-3-phosphate dehydrogenase, an RNA-binding protein), Pdc1p (pyruvate decarboxylase), and an unknown 35-kDa acidic protein. Copurification experiments demonstrated that Ssa1p bound to p33 replication protein in vivo, and surface plasmon resonance measurements with purified recombinant proteins confirmed this interaction in vitro. The double mutant strain (ssa1 ssa2) showed 75% reduction in viral RNA accumulation, whereas overexpression of either Ssa1p or Ssa2p stimulated viral RNA replication by approximately threefold. The activity of the purified CNV replicase correlated with viral RNA replication in the above-mentioned ssa1 ssa2 mutant and in the Ssa overexpression strains, suggesting that Ssa1/2p likely plays an important role in the assembly of the CNV replicase.
TL;DR: Carohydrate compositional as well as methylation analysis indicated that Asn29and Asn56 bear exclusively complex‐type oligosaccharide structures that are almost quantitatively α1‐6 fucosylated at the proximal N‐acetylglucosamine; ∼70% of these molecules contain a bisecting N‐ acetylglUCosamine.
Abstract: beta-Trace protein from pooled human CSF was purified to homogeneity. An apparent molecular mass of 23-29 kDa was determined for the polypeptide on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Amino-terminal sequencing of the polypeptide yielded the unique amino acid sequence APEAQVSVQPNFQQDKFLGRWFSA. Alignment of amino acid sequences obtained from tryptic peptides with the sequence previously deduced from a cDNA clone isolated by other investigators allowed the identification of beta-trace protein as prostaglandin D synthase [prostaglandin-H2 D-isomerase; (5Z, 13E)-(15S)-9 alpha, 11 alpha-epidioxy-15-hydroxyprosta-5,13-dienoate D-isomerase; EC 5.3.99.2]. A conservative amino acid exchange (Thr instead of Ser) was detected at amino acid position 154 of the beta-trace polypeptide chain in the corresponding tryptic peptide. The two N-glycosylation sites of the polypeptide were shown to be almost quantitatively occupied by carbohydrate. Carbohydrate compositional as well as methylation analysis indicated that Asn29 and Asn56 bear exclusively complex-type oligosaccharide structures (partially sialylated with alpha 2-3- and/or alpha 2-6-linked N-acetylneuraminic acid) that are almost quantitatively alpha 1-6 fucosylated at the proximal N-acetylglucosamine; approximately 70% of these molecules contain a bisecting N-acetylglucosamine. Agalacto structures as well as those with a peripheral fucose are also present.
TL;DR: The data suggest that sugar mobilization from glucose to coenzyme A and its acyl derivative is a collaboration between the cytosol and plastids and that temporal control of enzymes and pathways extends beyond transcription.
Abstract: Brassica napus (cultivar Reston) seed proteins were analyzed at 2, 3, 4, 5, and 6 weeks after flowering in biological quadruplicate using two-dimensional gel electrophoresis. Developmental expression profiles for 794 protein spot groups were established and hierarchical cluster analysis revealed 12 different expression trends. Tryptic peptides from each spot group were analyzed in duplicate using matrix-assisted laser desorption ionization time-of-flight mass spectrometry and liquid chromatography-tandem mass spectrometry. The identity of 517 spot groups was determined, representing 289 nonredundant proteins. These proteins were classified into 14 functional categories based upon the Arabidopsis (Arabidopsis thaliana) genome classification scheme. Energy and metabolism related proteins were highly represented in developing seed, accounting for 24.3% and 16.8% of the total proteins, respectively. Analysis of subclasses within the metabolism group revealed coordinated expression during seed filling. The influence of prominently expressed seed storage proteins on relative quantification data is discussed and an in silico subtraction method is presented. The preponderance of energy and metabolic proteins detected in this study provides an in-depth proteomic view on carbon assimilation in B. napus seed. These data suggest that sugar mobilization from glucose to coenzyme A and its acyl derivative is a collaboration between the cytosol and plastids and that temporal control of enzymes and pathways extends beyond transcription. This study provides a systematic analysis of metabolic processes operating in developing B. napus seed from the perspective of protein expression. Data generated from this study have been deposited into a web database (http://oilseedproteomics.missouri.edu) that is accessible to the public domain.
TL;DR: This work has shown that the gel-based and MuDPIT approaches have facilitated vital advances in the measurement of protein expression alterations in normal and disease phenotypic states, but the multiplexing fluorescent two-dimensional fluorescence difference gel electrophoresis capability minimizes spot pattern variability and the number of gels in an experiment while providing simple, accurate and reproducible spot matching.
Abstract: Quantitative proteomics is the workhorse of the modern proteomics initiative. The gel-based and MuDPIT approaches have facilitated vital advances in the measurement of protein expression alterations in normal and disease phenotypic states. The methodological advance in two-dimensional gel electrophoresis (2DGE) has been the multiplexing fluorescent two-dimensional fluorescence difference gel electrophoresis (2D-DIGE). 2D-DIGE is based on direct labeling of lysine groups on proteins with cyanine CyDye DIGE Fluor minimal dyes before isoelectric focusing, enabling the labeling of 2–3 samples with different dyes and electrophoresis of all the samples on the same 2D gel. This capability minimizes spot pattern variability and the number of gels in an experiment while providing simple, accurate and reproducible spot matching. This protocol can be completed in 3–5 weeks depending on the sample size of the experiment and the level of expertise of the investigator.
TL;DR: The analysis of posttranslational modifications revealed that none of the WSSV structural proteins was glycosylated and that VP28 and VP19 were threonine phosphorylated, which should provide an important reference for future molecular studies of WSSVs morphogenesis.
Abstract: White spot syndrome virus (WSSV) virions were purified from the tissues of infected Procambarus clarkii (crayfish) isolates. Pure WSSV preparations were subjected to Triton X-100 treatment to separate into the envelope and nucleocapsid fractions, which were subsequently separated by 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The major envelope and nucleocapsid proteins were identified by either matrix-assisted laser desorption ionization-time of flight mass spectrometry or defined antibody. A total of 30 structural proteins of WSSV were identified in this study; 22 of these were detected in the envelope fraction, 7 in the nucleocapsid fraction, and 1 in both the envelope and the nucleocapsid fractions. With the aid of specific antibodies, the localizations of eight proteins were further studied. The analysis of posttranslational modifications revealed that none of the WSSV structural proteins was glycosylated and that VP28 and VP19 were threonine phosphorylated. In addition, far-Western and coimmunoprecipitation experiments showed that VP28 interacted with both VP26 and VP24. In summary, the data obtained in this study should provide an important reference for future molecular studies of WSSV morphogenesis.
TL;DR: A novel 40-residue antimicrobial peptide, aurelin, exhibiting activity against Gram-positive and Gram-negative bacteria, was purified from the mesoglea of a scyphoid jellyfish Aurelia aurita by preparative gel electrophoresis and RP-HPLC.
TL;DR: The combination of multi‐dimensional liquid chromatography and two‐dimensional difference gel electrophoresis provides a valuable tool for serum proteomics in lung cancer.
Abstract: To investigate aberrant plasma proteins in lung cancer, we compared the proteomic profiles of serum from five lung cancer patients and from four healthy volunteers. Immuno-affinity chromatography was used to deplete highly abundant plasma proteins, and the resulting plasma samples were separated into eight fractions by anion-exchange chromatography. Quantitative protein profiles of the fractionated samples were generated by two-dimensional difference gel electrophoresis, in which the experimental samples and the internal control samples were labeled with different dyes and co-separated by two-dimensional polyacrylamide gel electrophoresis. This approach succeeded in resolving 3890 protein spots. For 364 of the protein spots, the expression level in lung cancer was more than twofold different from that in the healthy volunteers. These differences were statistically significant (Student's t-test, p-value less than 0.05). Mass spectrometric protein identification revealed that the 364 protein spots corresponded to 58 gene products, including the classical plasma proteins and the tissue-leakage proteins catalase, clusterin, ficolin, gelsolin, lumican, tetranectin, triosephosphate isomerase and vitronectin. The combination of multi-dimensional liquid chromatography and two-dimensional difference gel electrophoresis provides a valuable tool for serum proteomics in lung cancer.
TL;DR: A fluorescence-based approach using fluorescein-5-thiosemicarbazide (FTC) is adapted to quantify the global protein carbonyls as well as the carbonyl levels on individual proteins in the proteome and gives investigators the ability to identify potential target proteins that become oxidized under different pathological and physiological conditions.
TL;DR: Genomic DNA from methicillin-resistant Staphylococcus aureus isolates recovered from pigs and their caretakers proved resistant to SmaI digestion, leading to uninterpretable results in standard pulsed-field gel electrophoresis.
Abstract: Genomic DNA from methicillin-resistant Staphylococcus aureus isolates recovered from pigs and their caretakers proved resistant to SmaI digestion, leading to uninterpretable results in standard pulsed-field gel electrophoresis. This is the result of a yet unknown restriction/methylation system in the genus Staphylococcus with the recognition sequence CCNGG.
TL;DR: This review describes recent tendencies for metal-binding and metalloprotein analysis, emphasizing metal quantification in proteins through X-ray, atomic absorption, mass spectrometric techniques, and others.
TL;DR: 5'-Nucleotides of A and U with the phosphate activated with 1-methyladenine generate RNA oligomers containing 40-50 monomers in 1 day in reactions catalyzed by montmorillonite, the potential significance of these data to the origin and early evolution of life is discussed.
Abstract: 5‘-Nucleotides of A and U with the phosphate activated with 1-methyladenine generate RNA oligomers containing 40−50 monomers in 1 day in reactions catalyzed by montmorillonite. The corresponding monomers of C give oligomers that are 20−25-mers in length after a 9-day reaction. It was not possible to determine the chain lengths of the oligomers of G since they did not give well-defined bands on gel electrophoresis. Co-oligomers of A and U as well as A, U, G, and C were also prepared. The oligo(A)s formed were separated by gel electrophoresis, and the bands of the 7−39-mers were isolated, the 3‘,5‘-phosphodiester bonds were cleaved by RNase T2, and the terminal phosphate groups were cleaved with alkaline phosphatase. HPLC analysis revealed that the proportions of A5‘pp5‘A, A, A2‘pA, and A2‘pA2‘pA formed were almost the same for the long and shorter oligomers. A similar structure analysis performed on the oligo(U)s established that the proportions of U5‘pp5‘U, U, U2‘pU, U2‘pU2‘pU, U2‘pU2‘pU2‘pU, and U2‘pU2‘p...
TL;DR: In this study microwave technology was used to develop a fast protein preparation and enzymatic digestion method for protein mixtures, which enabled preparation and digestion ofprotein mixtures in solution or in gel in 6 or 25 min, respectively.
TL;DR: Flow cytometry analysis using fluorescence-activated cell sorting with fluorescein isothiocyanate-labeled annexin V demonstrated that the TTSS1-dependent cell death was by apoptosis, and cytotoxicity to HeLa cells was related to one of the newly identified secreted proteins encoded by VP1680.
Abstract: Vibrio parahaemolyticus, a gram-negative marine bacterium, is an important pathogen causing food-borne gastroenteritis or septicemia. Recent genome sequencing of the RIMD2210633 strain (a Kanagawa phenomenon-positive clinical isolate of serotype O3:K6) revealed that the strain has two sets of gene clusters that encode the type III secretion system (TTSS) apparatus. The first cluster, TTSS1, is located on the large chromosome, and the second, TTSS2, is on the small chromosome. Previously, we reported that TTSS1 is involved in the cytotoxicity of the RIMD2210633 strain against HeLa cells. Here, we analyzed proteins secreted via the TTSS apparatus encoded by TTSS1 by using two-dimensional gel electrophoresis and identified the proteins encoded by genes VP1680, VP1686, and VPA450. To investigate the roles of those secreted proteins, we constructed and analyzed a series of deletion mutants. Flow cytometry analysis using fluorescence-activated cell sorting with fluorescein isothiocyanate-labeled annexin V demonstrated that the TTSS1-dependent cell death was by apoptosis. The cytotoxicity to HeLa cells was related to one of the newly identified secreted proteins encoded by VP1680. Adenylate cyclase fusion protein studies proved that the newly identified secreted proteins were translocated into HeLa cells. Thus, these appear to be the TTSS effector proteins in V. parahaemolyticus.
TL;DR: There were only minor changes in the major proteins secreted in the epididymal activity in humans, and it is suggested that this specificity might be related to the difference between species in the location of the epidIDymis where sperm become fertile.
TL;DR: QD-decorated plasmid DNA was utilized for the first time for long-term intracellular and intranuclear tracking studies and was capable of expressing the reporter protein, enhanced green fluorescent protein, following transfection in Chinese hamster ovary (CHO-K1) cells with an efficiency of ca.
TL;DR: By combining subcellular fractionation with high resolution separation and comprehensive identification of proteins, this work has begun to elucidate zymogen granule functions through proteomic and subsequent functional analysis of its membrane components.
TL;DR: This protocol describes the separation of proteins by SDS-polyacrylamide gel electrophoresis with the aim of estimating the molecular weight of the polypeptide chain(s) by using markers of known molecular weight.
Abstract: INTRODUCTIONThis protocol describes the separation of proteins by SDS-polyacrylamide gel electrophoresis. SDS is used with a reducing agent and heat to dissociate the proteins. SDS-polypeptide complexes form and migrate through the gels according to the size of the polypeptide. By using markers of known molecular weight, the molecular weight of the polypeptide chain(s) can be estimated.
TL;DR: Results showed that proteins induced by heavy metal exposure are principally involved in oxidative stress tolerance or in other stress pathways, which could lead to a better understanding of the mechanisms involved in cell protection and tolerance.