Isolation, Library Preparation, and Bioinformatic Analysis of Historical and Ancient Plant DNA

The ability to sequence DNA retrieved from ancient and historical material plays a crucial role in reinforcing evolutionary and anthropological inference. While the focus of the field is largely on analyzing DNA from ancient hominids and other animals, we have also learned from plant ancient DNA (aDNA), in particular, about human farming practices, crop domestication, environment management, species invasion, and adaptation to various environmental conditions. In the following protocols, we outline best practices for plant aDNA isolation, preparation for sequencing, bioinformatic processing, and authentication. We describe the process all the way from processing of archaeological or historical plant material to characterizing and authenticating sequencing reads. In alternative protocols, we include modifications to this process that are tailored to strongly degraded DNA. Throughout, we stress the importance of precautionary measures to successfully analyze aDNA. Finally, we discuss the evolution of the archaeogenomics field and the development of new methods, which both shaped this protocol. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Isolation of aDNA Alternate Protocol 1: Isolation of ultra-short DNA (Dabney modification) Support Protocol 1: Preparation of PTB-based mix Support Protocol 2: Preparation of binding buffer Basic Protocol 2: Preparation of genomic libraries Alternate Protocol 2: Preparation of genomic libraries with uracil removal Basic Protocol 3: Bioinformatic processing and authentication of aDNA

The Arabidopsis thaliana F-box gene HAWAIIAN SKIRT is a new player in the microRNA pathway

In Arabidopsis, the F-box HAWAIIAN SKIRT (HWS) protein is important for organ growth. Loss of function of HWS exhibits pleiotropic phenotypes including sepal fusion. To dissect the HWS role, we EMS-mutagenized hws-1 seeds and screened for mutations that suppress hws-1 associated phenotypes. We identified shs-2 and shs-3 (suppressor of hws-2 and 3) mutants in which the sepal fusion phenotype of hws-1 was suppressed. shs-2 and shs-3 (renamed hst-23/hws-1 and hst-24/hws-1) carry transition mutations that result in premature terminations in the plant homolog of Exportin-5 HASTY (HST), known to be important in miRNA biogenesis, function and transport. Genetic crosses between hws-1 and mutant lines for genes in the miRNA pathway, also suppress the phenotypes associated with HWS loss of function, corroborating epistatic relations between the miRNA pathway genes and HWS. In agreement with these data, accumulation of miRNA is modified in HWS loss or gain of function mutants. Our data propose HWS as a new player in the miRNA pathway, important for plant growth

Sex- and age-related reference values in cardiology, with annotations and guidelines for interpretation

The definition of “abnormal” in clinical sciences is often based on so-called reference values which point to a range that experts by some sort of consensus consider as normal when looking at biological variables. Such a level is commonly calculated by taking (twice) the standard deviation from the mean, or considering certain percentiles. The suspicion or even confirmation of a disease is then established by demonstrating that the value measured exceeds the upper or lower reference value. As is often the case, the measurement accuracy may depend on the conditions and specific method employed to collect and analyze data. This implies that, for example, data assessed by 2D echocardiography possibly differ from those obtained by MRI and therefore require modality-specific reference values. In this review we summarize reference values for the electrocardiogram, cardiac compartmental volumes, and arterial vessel size in males and females for various age groups. These values may further depend on other variables such as body size, physical training status, and ethnicity. Additional variables relevant for cardiology such as those referring to the microcirculation and biomarkers are only mentioned with reference to the pertinent literature. In general, the sex- and age-specific differences observed are often remarkable and warrant consideration in clinical practice and basic biomedical sciences