Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.64 (proteinase K)
 4,071 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Sequences of hepatitis B virus DNA were detected specifically in the sera of infected individuals by a non-radioactive riboprobe nucleic acid hybridization assay. The nucleic acids contained in serum specimens were prepared by an overnight proteinase K-/SDS-digest, phenol/chloroform-extraction and ethanol-precipitation, and immobilized on nylon membranes by the dot-blot technique. Digoxigenin-labelled 1.4 kb RNA-probes were generated by the phage sp 6 RNA-polymerase from a linearized HBV DNA transcription template containing the appropriate promoter. Hybridized probes were detected by alkaline phosphatase-conjugated sheep anti-digoxigenin Fab-fragments, and 5-bromo-4-chloro-3-indolylphosphate (BCIP) and nitroblue tetrazolium salt (NBT) as chromogenic substrates for the subsequent ELISA procedure. With a detection limit of 0.5-1.0 pg HBV DNA and a high specificity, the results were comparable to those obtained by the standard assay employing radioactively labeled DNA-probes.
 ...
PMID:A nonradioactive riboprobe assay for the detection of hepatitis B virus DNA in human sera. 208

A method is described for the rapid isolation of chromosomal deoxyribonucleic acid from species of the genus Mycoplasma. The method involves incubation of washed cells at elevated temperature in the presence of an ionic detergent, chelating agents, and proteinase K prior to the removal of residual protein and ribonucleic acid with ribonuclease and chloroform. It results in a good yield of high molecular weight material that is shown to be free of endogenous nuclease and substantially free of protein or ribonucleic acid contamination without the use of phenol. The isolated DNA is shown to be an excellent substrate for restriction endonuclease digestion and ligation with T4 DNA ligase.
 ...
PMID:An improved method for the rapid isolation of chromosomal DNA from Mycoplasma spp. 218 71

Modification of the alkaline lysis at elevated temperature technique is proposed isolation of plasmid DNA from lactobacilli. Modification consists of colorimetric control of culture phase during the biomass growth, pH control at the probes treatment with lysozyme and alkaline solution of natrium dodecylsulfate by adding the indicator bromcrezolpurple into the medium for biomass growth. The high concentration of lysozyme is used (10 mkg.ml-1). Lactobacilli are lysed at 2 min incubations of the probes with the lytic solution in the boiling water bath. The treatment of the probes by proteinase K, by the mixture of chloroform:phenol:isoamyl spirit (25:24:1 vol/vol/vol) and by diethylpirocarbonate increased considerably the quality of the obtained DNA preparations. The modified technique is suitable for isolation of the plasmid DNA from lactobacilli of different species, enterococci, streptococci and other lactic bacteria. The connection of antibiotic resistance marker and the plasmid profile of lactobacilli under different conditions with the presence of the plasmid DNA- protein complex is discussed.
 ...
PMID:[Optimization of the method of isolation of microamounts of plasmid DNA from lactobacilli]. 236

In order to establish a routine procedure for isolating undegraded RNA from small amounts of rat and human brain tissue, several techniques were investigated. Initial studies demonstrated that undegraded RNA could not be reproducibly isolated from milligram amounts of brain tissue homogenized in an aqueous medium. Several isolation techniques utilizing tissue homogenization in the denaturing agent guanidinium chloride were compared. This method of homogenization, followed by sedimentation of RNA through cesium chloride, resulted in good yields of undegraded translationally active RNA. A maximum of 6 RNA samples could be processed simultaneously. In contrast, when homogenization in guanidinium chloride was followed by repeated guanidinium chloride-ethanol precipitations many samples could be processed simultaneously. The resulting RNA yields were low. The introduction of several modifications in the guanidinium chloride-ethanol precipitation technique resulted in a high yield of undegraded translationally active RNA. DNA was removed by two guanidinium-ethanol precipitations. Residual protein was digested with proteinase K. RNA was precipitated after extraction with phenol-chloroform-isoamyl alcohol. This refined procedure allows the recovery, in high yields, of translationally active undegraded RNA which is both DNA and protein free. Thirty-six samples can be processed in one day.
 ...
PMID:A rapid microprocedure for isolating RNA from multiple samples of human and rat brain. 241 66

We have developed a new, rapid method for the extraction of human genomic DNA from whole blood samples. Traditionally, genomic DNA has been extracted from blood by overnight proteinase K digestion of lysed peripheral lymphocytes followed by phenol/chloroform extraction. In addition to being time consuming, the use of phenol involves inherent risks due to the toxic nature of the reagent. Our method for the extraction of DNA from whole blood uses sodium perchlorate and chloroform instead of phenol with a significant time savings realized as well as fewer hazards to the technician. Furthermore, DNA prepared by this new method is an excellent substrate for restriction endonuclease digestion and Southern hybridization analysis.
 ...
PMID:Purification of human genomic DNA from whole blood using sodium perchlorate in place of phenol. 255 54

Frozen sections of human gingiva and skin, fixed in acetone, were subjected to limited enzyme digestion (neuraminidase, proteinase K, trypsin) or, respectively, the application of solvents (chloroform/methanol, triton X-100) to allow a partial characterization of epithelial lectin binding sites. Gingiva differs from normal skin in that more conA-binding glycolipids are present in the lower cell layers. In the upper layers conA-fixing glycoproteins are prevailing. Psoriatic foci regularly exhibit an increased presence of conA-binding glycolipids. Gingiva and normal skin have some common features in the behavior of the lectin binding sites of HPA, WGA and UEA I. Analogies in the binding pattern of conA and UEA I in gingival tissue and in psoriatic foci are thus due to different lectin receptors.
 ...
PMID:[Partial characterization of the epithelial lectin binding sites of human gingiva and skin]. 259 11

T lymphocyte-mediated immunity is important for resistance to Francisella tularensis. To characterize the specificity of this immunity, we used membrane proteins and two lipopolysaccharide (LPS) preparations. Both membrane proteins were heat-modifiable, as indicated by their migration in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). One had an apparent molecular mass (Mm) of 120 kilodaltons (kDa) when solubilized in the SDS buffer at room temperature, but 17 kDa after heating. The respective values for the other protein were 35 kDa before and 40 kDa after heating. Both proteins were purified by a preparative SDS-PAGE. The LPS-containing preparations were isolated by aqueous phenol (WP) or PCP (phenol-chloroform-petroleum ether) extraction (LPS-R), and rendered protein-free by treatment with proteinase K. Lymphocytes from nine subjects immunized with a live tularemia vaccine from one to three years earlier responded specifically to both an F. tularensis whole cell antigen and the 17 kDa protein in the lymphocyte blast transformation test. By contrast, the 40 kDa protein and the two LPS preparations did not stimulate any detectable lymphocyte proliferation.
 ...
PMID:Membrane proteins of Francisella tularensis LVS differ in ability to induce proliferation of lymphocytes from tularemia-vaccinated individuals. 262 30

A method is described which facilitates the rapid purification of high molecular weight chromosomal DNA from gram positive and gram negative bacteria grown on solid media. A total of 32 reference strains and fresh isolates were examined in this study. The purification procedure involved lysis of cells with SDS in the presence of proteinase K, followed by removal of cellular polysaccharides and proteins with hexadecyltrimethyl ammonium bromide (CTAB) and phenol:chloroform:isoamyl alcohol. Preparations were incubated with RNase and, after removal of the enzyme, DNA was precipitated with ethanol. Several hundred micrograms of DNA could be prepared within 5 h from cells grown on 1-2 agar plates. None of the final preparations contained RNA; protein was detected in 12/32 preparations. The resultant DNA proved suitable for restriction enzyme digestion and biotin-labelling by a random primer technique. DNA probes constructed from these preparations were capable of detecting 100 pg of homologous target DNA fixed to nitrocellulose. Cross reactions between closely related species displayed weaker signal intensities than, and, thus, were easily distinguished from, true positive reactions between homologous species. DNA obtained by this procedure may also be suitable for DNA-DNA homology studies, recombinant DNA experiments and molecular fingerprinting.
 ...
PMID:Rapid method for the purification of DNA from subgingival microorganisms. 262 68

A simplified method for preparing genomic DNA at and human brain has been established. The procedure includes the following steps: (1) mincing and homogenizing in SSC-EDTA, (2) solubilizing in high concentration detergent (2% SDS or Triton X-100), (3) digesting with proteinase K and RNase A, (4) extracting with phenol-chloroform, (5) finally dialyzing the sample against TE. This method could result in a threefold increase in yield with advantages of simplification and time-saving. The DNA obtained by this method is of a size over 30 kd, and is suitable for the molecular biological and molecular genetical study of brain.
 ...
PMID:[A molecular biology technique for the brain--preparation of genomic DNA from the brain]. 263 87

Modification of Autographa californica multicapsid nuclear polyhedrosis virus extracellular virion polypeptides with fatty acid was investigated. Fatty acid-derived radioactivity was incorporated into gp67, the predominant virion envelope glycoprotein, during metabolic labeling using [3H]palmitic acid or [3H]myristic acid. The acyl moiety resisted extraction with chloroform-methanol but was released by mild alkali or hydroxylamine treatment, features characteristic of an ester-type linkage. Palmitic acid was identified as the fatty acid liberated during acid hydrolysis of purified [3H]palmitic acid and [3H]myristic acid-labeled gp67. Acyl peptides were protected during proteinase K digestion of intact virions but they were larger (Mr 34.3K and 31.8K) than expected on the basis of membrane protection alone. Carbohydrate may have played a role in determining the protease resistance of the observed fragments. Potential acylation sites on gp67 within or adjacent to the C-terminal hydrophobic region are discussed with respect to the predicted amino acid sequence.
 ...
PMID:Fatty acid acylation of the 67K envelope glycoprotein of a baculovirus: Autographa californica nuclear polyhedrosis virus. 267 65


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>