ABSTRACT
The restriction endonuclease BliAI, an isoschizomer of ClaI, which recognizes the sequence 5'- AT¼CGAT - 3', was purified from a natural isolate identified as Bacillus licheniformis. The restriction endonuclease was isolated from cell extracts using single-step purification by phosphocellulose column chromatography. The restriction endonuclease is active at 37°C and over a wide range of pH and salt concentration. The molecular weight of the purified restriction enzyme is consistent with a value of 39 kDa.
A endonuclease de restrição BliAI, um isoesquisômero de ClaI, que reconhece a seqüência 5'- AT¼CGAT - 3', foi purificada de um isolado natural identificado como Bacillus licheniformis. A endonuclease de restrição em questão foi isolada a partir de um extrato celular em um único passo cromatográfico utilizando uma coluna contendo a resina fosfocelulose. A endonuclease de restrição é ativa à 37°C e em uma ampla escala de pH e concentração de sais. O peso molecular da enzima purificada corresponde a um valor de kDa 39.
Subject(s)
Bacillus , Clinical Enzyme Tests , Deoxyribonucleases, Type I Site-Specific , Endonucleases , In Vitro Techniques , Chromatography , Molecular Weight , Water SamplesABSTRACT
<p><b>OBJECTIVE</b>To map the regulatory domain of Escherichia coli T-protein.</p><p><b>METHODS</b>Fragmentation cloning was employed in cloning of 11 fragments from T-protein. The regulatory activity of each fragment was determined respectively.</p><p><b>RESULTS</b>The regulatory domain of T-protein was located in the C-terminal 270 amino acids, which was the same location as PDH domain.</p><p><b>CONCLUSION</b>T-protein has no independent regulatory domain.</p>
Subject(s)
Cloning, Molecular , Deoxyribonucleases, Type I Site-Specific , Chemistry , Genetics , Escherichia coli Proteins , Chemistry , Genetics , Nucleic Acid Conformation , Peptide Fragments , Chemistry , Genetics , Protein Binding , Genetics , Regulatory Elements, Transcriptional , GeneticsABSTRACT
Background. The MPS-I is an autosomal recessive disorder caused by mutations in the IDUA gene that induce to a deficiency of glycosidase Ó-L-iduronidase that is required for degradation of heparan and dermatan sulfate. This disorder expresses a wide range of clinical symptoms. Methods. Kpnl (k) and VNTR (V) intragenic polymorphisms at the IDUA gene were studied in mestizo and Huichol Indian Mexican populations as well in 13 MPS-I patients. Data from Australian normal and MPS-I (2-4) individuals were also studied. Results. Genotypes for IDUA K and V sites in Mexicans were in agreement with hardy-Weinberg expectations, except for stie K in Huichols, Individually, allele frequency distributions were different (p< 0.05) in the two normal groups for the V site. K-V haplotype frequency distributions (HFDs) in these two normal groups were also different as compared with normal Australians. In Mexican MPS-I patients, HFD was different (p <0.05) with or MPS-I Australians. This can be taken as evidence of linkage disequilibrium between K-V polymorphism and MPS-I gene mutation(s) at the IDUA region. A similar finding was reported. However, disequilibrium in Mexicans was determined by haplotypes different from those in Australia. In Mexican MPS-I patients, haplotype K2-V1 is increased and K1-V3 decreades with respect to the Mexican mestizo (p < 0.05), while in Australians, MPS-I patients had an increase of haplotypes K2-V2 and K1-V2 with respect to expected frequency. Conclusions. The similar HFD between Mexican and australian MPS-I patients suggests a common genetic origin, that MPS-I mutations were introduced to Mexico by Spaniards, and that such mutations predate the dispersion between Mexican and Australian Caucasian ancestors. The differences in disequilibrium are explained rather by genetic drift
Subject(s)
Deoxyribonucleases, Type I Site-Specific , Haplotypes , Linkage Disequilibrium , Mucopolysaccharidosis I/ethnology , Mucopolysaccharidosis I/genetics , Base Sequence , MexicoABSTRACT
En el presente trabajo se describen parte de las técnicas de biología molecular que se utilizan en medicina. Actualmente las principales aplicaciones de estas técnicas son en el diagnóstico de enfermedades hereditarias, infecciosas y neoplásicas, así como en estudios de regulación de la expresión génica y en terapia con proteínas recombinantes. Con la aplicación de las técnicas de biología molecular en diversas áreas de la medicina, se abre también la posibilidad de poder realizar terapia génica en humanos