The impact of pre-analytical processing on staining quality for H&E, dual hapten, dual color in situ hybridization and fluorescent in situ hybridization assays.

With the advent of personalized medicine, anatomic pathology-based molecular assays, including in situ hybridization (ISH) and mRNA detection tests, are performed routinely in many laboratories and have increased in their clinical importance and complexity. These assays require appropriately fixed tissue samples that preserve both nucleic acid targets and histomorphology to ensure reliable test results for determining patient treatment options. However, all aspects of tissue processing, including time until tissue fixation, type of fixative, duration of fixation, post-fixation treatments, and sectioning of the sample, impact the staining results.

Early interrogation and recognition of DNA sequence by indirect readout.

Control of replication, transcription, recombination and repair requires proteins capable of finding particular DNA sequences in a background of a large excess of nonspecific sequences. Such recognition can involve direct readout, with direct contacts to the bases of DNA, or in some cases through the less well-characterized indirect readout mechanisms. In order to measure the relative contributions of direct and indirect readout by a sequence specific endonuclease, HincII, a mutant enzyme deficient in a direct contact, was characterized, and surprisingly showed no loss of sequence specificity.

DNA distortion and specificity in a sequence-specific endonuclease.

Five new structures of the Q138F HincII enzyme bound to a total of three different DNA sequences and three different metal ions (Ca(2+), Mg(2+), and Mn(2+)) are presented. While previous structures were produced from soaking Ca(2+) into preformed Q138F HincII/DNA crystals, the new structures are derived from cocrystallization with Ca(2+), Mg(2+), or Mn(2+). The Mn(2)(+)-bound structure provides the first view of a product complex of Q138F HincII with cleaved DNA.

Cooperative DNA binding by CI repressor is dispensable in a phage lambda variant.

Complex gene regulatory circuits contain many interacting components. In principle, all of these components and interactions may be essential to the function of the circuit. Alternatively, some of them may be refinements to a simpler version of the circuit that improve its fitness.

Conservation and diversity in the immunity regions of wild phages with the immunity specificity of phage lambda.

The gene regulatory circuitry of phage lambda is among the best-understood circuits. Much of the circuitry centres around the immunity region, which includes genes for two repressors, CI and Cro, and their cis-acting sites. Related phages, termed lambdoid phages, have different immunity regions, but similar regulatory circuitry and genome organization to that of lambda, and show a mosaic organization, arising by recombination between lambdoid phages.