Strand separation

There is still a controversy over the mechanism of promoter DNA strand separation upon open transcription complex (RPo) formation by Escheri-chia coli RNA polymerase: is it a single or a stepwise process controlled by Mg 2+ ions and temperature? To resolve this question, the kinetics of pseudo-first-order oxidation of thymine residues by KMnO4 in the )11…+2 DNA region of RPo at thekPRpromoter was examined under single-hit conditions as a function of temperature (13–37
C) in the absence or presence of 10 mmMgCl2....

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DNA helicases are molecular
motor enzymes that use the energy of NTP hydrolysis to separate transiently energetic-ally stable duplex DNA into single strands. They are there-fore essential in nearly all DNA metabolic transactions. They act as essential molecular tools for the cellular machinery. Since the discovery of the first DNA helicase in Escherichia coliin1976, several havebeen isolated fromboth prokaryotic and eukaryotic systems.

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The occurrence of DNA double-strand breaks in the nucleus provokes in its structural organization a large-scale alteration whose molecular basis is still mostly unclear. Here, we show that double-strand breaks trigger pref-erential assembly of nucleoproteins in human cellular fractions and that they mediate the separation of large protein–DNA aggregates from aque-ous solution.

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Automated DNA sequencing is a core research tool used by almost every research biochemistry lab. It is used to determine the sequence of DNA, or the genetic code, that serves as the blueprint of life for every organism on Earth.

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Concept of hybrid molecules: When double strand DNA is steamed to a temperature exceed the melting temperature (Tm), it will separate into 2 single strands DNA due to breaks of H bonds. If the reaction temperature is then decreased slowly plus other appropriate experimental conditions, these ssDNA will pair again. This phenomenon is called the hybridization of molecules. Characteristic of the hybrid molecules: Specificity, the re-pairing occurs only between two complementary sequences.

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More discrete sequence alterations rely heavily on the use of the PCR, which allows rapid gene amplification and analysis. Moreover, PCR makes it possible to perform genetic testing and mutational analysis with small amounts of DNA extracted from leukocytes or even from single cells, buccal cells, or hair roots. Screening for point mutations can be performed by numerous methods (Table 62-9); most are based on the recognition of mismatches between nucleic acid duplexes, electrophoretic separation of single- or double-stranded DNA, or sequencing of DNA fragments amplified by PCR.

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