Signal and noise in bridging PCR

BACKGROUND: In a variant of the standard PCR reaction termed bridging, or jumping, PCR the primer-bound sequences are originally on separate template molecules. Bridging can occur if, and only if, the templates contain a region of sequence similarity. A 3' end of synthesis in one round of synthesis that terminates in this region of similarity can prime on the other. In principle, Bridging PCR (BPCR) can detect a subpopulation of one template that terminates synthesis in the region of sequence shared by the other template. This study considers the sensitivity and noise of BPCR as a quantitative assay for backbone interruptions. Bridging synthesis is also important to some methods for computing with DNA. RESULTS: In this study, BPCR was tested over a 328 base pair segment of the E. coli lac operon and a signal to noise ratio (S/N) of approximately 10 was obtained under normal PCR conditions with Taq polymerase. With special precautions in the case of Taq or by using the Stoffel fragment the S/N was improved to 100, i.e. 1 part of cut input DNA yielded the same output as 100 parts of intact input DNA. CONCLUSIONS: In the E. coli lac operator region studied here, depending on details of protocol, between 3 and 30% per kilobase of final PCR product resulted from bridging. Other systems are expected to differ in the proportion of product that is bridged consequent to PCR protocol and the sequence analyzed. In many cases physical bridging during PCR will have no informational consequence because the bridged templates are of identical sequence, but in a number of special cases bridging creates, or, destroys, information

Method for Linking a Synthesized Protein to Its mRNA-DNA Complex

A nascent protein remains in a complex with its ribosome and mRNA if the stop codon is deleted from the mRNA. In the same manner, mRNA forms a stable complex with DNA if the transcription termination is blocked. In principle, if both mRNA translation and DNA transcription termination are prevented, the protein should stay in a complex with its mRNA and DNA. A method is designed to test these possibilities. Using an immobilized luciferase gene sequence, a functional luciferase protein is produced that remains associated through its mRNA with its DNA, confirming the feasibility of the proposed scheme. It has potential application for in vitro synthesis of proteins and protein micro-arrays

Characteristic slip for five great earthquakes along the Fuyun fault in China

International audienceThe seismic hazard associated with an individual fault can be assessed from the distributions of slip and recurrence times of earthquakes. However, seismic cycle models that aim to predict rupture lengths and fault displacements of successive earthquakes on one fault remain poorly validated. It is therefore unknown whether individual fault segments rupture independently, producing earthquakes with a diverse range of magnitudes and recurrence times, or slip by characteristic amounts, with characteristic magnitudes. Here we use high-resolution satellite data to document the horizontal offsets of stream channels and terraces created by strike-slip motion on the Fuyun fault, Xinjiang, China, during five historical earthquakes. We find that the Ms 7.9 11 August 1931 earthquake produced a surface rupture with a length of 160km, dispersed over three different fault segments. The 290 measured stream channel and terrace offsets record an average slip of 6.3m. We use the degree of preservation of geomorphological markers to assign relative ages to individual fault offsets and identify at least four distinct older earthquakes. We find that these older earthquakes also produced fault offsets with a similar distribution to the 1931 earthquake. As the slip distributions during five successive earthquakes were so similar, we conclude that ruptures on the Fuyun fault obey a characteristic slip model