A novel strategy for heat-mediated activation of recombinant Taq DNA polymerase is described. A serum albumin binding protein tag is used to affinity-immobilize an E. coli-expressed Taq DNA polymerase fusion protein onto a solid support coated with human serum albumin (HSA). Analysis of heat-mediated elution showed that elevated temperatures (> 70 degrees C) were required to significantly relea...

We report here the rational design and optimization of an antibody-responsive, DNA-based device that enables communication between pairs otherwise non-interacting proteins. The is designed to recognize bind a specific antibody and, in response, undergo conformational change leads release DNA strand, termed “translator,” regulates activity downstream target protein. As proof principle, we demons...

Polymerase Chain Reaction (PCR) merupakan uji diagnostik yang sangat sensitif. Enzim Taq salah satu komponen paling penting dalam pengujian (PCR). taq polymerase harganya mahal. Penggunaan pada PCR kurang efisien. Untuk mengetahui efisiensi penggunaan enzim dan volume terkecil berapa masih dapat digunakan metode PCR, maka dilakukanlah penelitian dengan membuat lebih kecil (5µl, 10 µl, 25 µl) da...

For the detection of RNA transcripts by RT-PCR, prior removal of genomic DNA must be performed. To remove genomic DNA, RNA is often prepared by DNase I digestion following phenol extraction. Recently, one-tube or onebuffer systems of RT-PCR were developed to prevent loss of RNA and to reduce the risk of contamination (2,6). In these methods, DNase I is added before RT. Taq DNA polymerase prepar...

AIM To investigate the dose-response effect of humic acid (HA) on the quantitative real time polymerase chain reaction (QRT-PCR) inhibition and the efficiency of Taq polymerase increment in preventing inhibition by HA in DNA extracted from ancient bones. METHODS DNA was isolated from bone samples and DNA quantification was conducted with the real-time 5' exonuclease detection assay (TaqMan), ...

The preparation of cDNA libraries usually involves multiple steps, such as isolation of poly A mRNA, synthesis of cDNA, cloning of cDNA into plasmid, and amplification of cloned plasmid. The last step, the amplification of insert DNA fragment after cloned in dephosphorylated plasmid vector, is done in either of two ways. The recombinant DNA can be extracted after amplification in host cells, or...

DNA binding of the Type 1 DNA polymerase from Thermus aquaticus (Taq polymerase) and its Klentaq large fragment domain have been studied as a function of temperature. Equilibrium binding assays were performed from 5 to 70 degrees C using a fluorescence anisotropy assay and from 10 to 60 degrees C using isothermal titration calorimetry. In contrast to the usual behavior of thermophilic proteins ...

Important in understanding the regulation of gene transcription is the elucidation of specific protein-DNA interactions that occur in vivo. One strategy for such in vivo footprinting involves the treatment of whole cells with dimethyl sulphate (DMS), which leads to methylation of guanine residues in DNA at the N7 position (1, 2). This N7 atom lies in the major groove and its susceptibility to m...