Establishing broad generality of DNA catalysts for site-specific hydrolysis of single-stranded DNA

We recently reported that a DNA catalyst (deoxyribozyme) can site-specifically hydrolyze DNA on the minutes time scale. Sequence specificity is provided by Watson-Crick base pairing between the DNA substrate and two oligonucleotide binding arms that flank the 40-nt catalytic region of the deoxyribozyme. The DNA catalyst from our recent in vitro selection effort, 10MD5, can cleave a single-stranded DNA substrate sequence with the aid of Zn2+ and Mn2+ cofactors, as long as the substrate cleavage site encompasses the four particular nucleotides ATG^T. Thus, 10MD5 can cleave only 1 out of every 256 (44) arbitrarily chosen DNA sites, which is rather poor substrate sequence tolerance. In this study, we demonstrated substantially broader generality of deoxyribozymes for site-specific DNA hydrolysis. New selection experiments were performed, revealing the optimality of presenting only one or two unpaired DNA substrate nucleotides to the N40 DNA catalytic region. Comprehensive selections were then performed, including in some cases a key selection pressure to cleave the substrate at a predetermined site. These efforts led to identification of numerous new DNA-hydrolyzing deoxyribozymes, many of which require merely two particular nucleotide identities at the cleavage site (e.g. T^G), while retaining Watson-Crick sequence generality beyond those nucleotides along with useful cleavage rates. These findings establish experimentally that broadly sequence-tolerant and site-specific deoxyribozymes are readily identified for hydrolysis of single-stranded DNA

Global or regional? Constraining the origins of the middle Bambuí carbon cycle anomaly in Brazil

The Ediacaran-Cambrian Bambuí Group in Brazil records an anomalously positive excursion in carbonate carbon isotopes (δ13Ccarb) with a sustained plateau of ca. +15‰ (aka the Middle Bambuí Excursion–MIBE). Considering that the δ13Ccarb signals in Ediacaran-Cambrian seawaters do not typically exceed +6‰, the MIBE therefore represents a profound carbon cycle anomaly in Earth's history. Although intensive studies have been done on the Bambuí Group, origins of the MIBE remain enigmatic. In order to better constrain the biogeochemical carbon and sulfur cycles during the MIBE, high-resolution chemostratigraphic analysis was conducted for both the plateau (i.e., Lagoa do Jacaré Formation) and the recovery part (i.e., lower Serra da Saudade Formation) of the MIBE. Chemostratigraphic profiles reveal remarkably different values in δ13Ccarb, δ13Corg, δ18Ocarb, and δ34Spyrite between these two studied MIBE intervals. The new data show that the plateau of the MIBE is characterized by coupled higher δ13Ccarb, higher δ13Corg, and higher δ34Spyrite signals compared with the recovery part of the MIBE. Based on multiple lines of sedimentological, geochemical, and model evidence, we propose that the possibilities of enhanced organic carbon burial and porewater methanogenesis are insufficient to explain the MIBE. Instead, local or regional controlling factors, including water-column methanogenesis, low-sulfate conditions, and enhanced carbonate recycling in a restricted basin may have played a role, independently or in unison, in generating this profound positive δ13Ccarb excursion. Therefore, the MIBE may reflect a regional event, instead of a global carbon cycle anomaly. We caution against the use of the MIBE in chemostratigraphic correlations on a global scale or any other attempt to infer global carbon cycling at that time. The biogeochemical landscape of the late Ediacaran-Cambrian basins and ocean margins may be more heterogeneous than previously thought