Improved mammalian family phylogeny using gap-rare multiple sequence alignment: A timetree of extant placentals and marsupials.

The timing of mammalian diversification in relation to the Cretaceous-Paleogene (KPg) mass extinction continues to be a subject of substantial debate. Previous studies have either focused on limited taxonomic samples with available whole-genome data or relied on short sequence alignments coupled with extensive species samples. In the present study, we improved an existing dataset from the landmark study of Meredith et al. (2011) by filling in missing fragments and further generated another dataset containing 120 taxa and 98 exonic markers. Using these two datasets, we then constructed phylogenies for extant mammalian families, providing improved resolution of many conflicting relationships. Moreover, the timetrees generated, which were calibrated using appropriate molecular clock models and multiple fossil records, indicated that the interordinal diversification of placental mammals initiated before the Late Cretaceous period. Additionally, intraordinal diversification of both extant placental and marsupial lineages accelerated after the KPg boundary, supporting the hypothesis that the availability of numerous vacant ecological niches subsequent to the mass extinction event facilitated rapid diversification. Thus, our results support a scenario of placental radiation characterized by both basal cladogenesis and active interordinal divergences spanning from the Late Cretaceous into the Paleogene.

The eggshell-matrix protein gene OC-17 is functionally lost in the viviparous Chinese crocodile lizard.

Thickness reduction or loss of the calcareous eggshell is one of major phenotypic changes in the transition from oviparity to viviparity. Whether the reduction of eggshells in viviparous squamates is associated with specific gene losses is unknown. Taking advantage of a newly generated high-quality genome of the viviparous Chinese crocodile lizard (Shinisaurus crocodilurus), we found that ovocleidin-17 gene (OC-17), which encodes an eggshell matrix protein that is essential for calcium deposition in eggshells, is not intact in the crocodile lizard genome. Only OC-17 transcript fragments were found in the oviduct transcriptome, and no OC-17 peptides were identified in the eggshell proteome of crocodile lizards. In contrast, OC-17 was present in the eggshells of the oviparous Mongolia racerunner (Eremias argus). Although the loss of OC-17 is not common in viviparous species, viviparous squamates show fewer intact eggshell-specific proteins than oviparous squamates. Our study implies that functional loss of eggshell-matrix protein genes may be involved in the reduction of eggshells during the transition from oviparity to viviparity in the crocodile lizard.

Directing group strategies in rhodium-catalyzed C-H amination.

Construction of a carbon-nitrogen bond is one of the most prevalent operations in nature and organic synthesis. The resulting amino compounds are privileged structural fragments in natural products, pharmaceutical drugs, and functional materials. With the rapid advancement of C-H bond activation, directing-group strategies in C-H amination catalyzed by rhodium have emerged. This reaction approach considerably enhances the step economy and atom economy of the reaction, and it complies with green chemistry and atom economy. The reactivity and selectivity of chelation-assisted rhodium-catalyzed C-H amination are discussed in relation to the types of aminating reagent, as well as the challenges and future development prospects in this field.

Population genomics reveals that natural variation in PRDM16 contributes to cold tolerance in domestic cattle.

Environmental temperature serves as a major driver of adaptive changes in wild organisms. To discover the mechanisms underpinning cold tolerance in domestic animals, we sequenced the genomes of 28 cattle from warm and cold areas across China. By characterizing the population structure and demographic history, we identified two genetic clusters, i.e., northern and southern groups, as well as a common historic population peak at 30 kilo years ago. Genomic scan of cold-tolerant breeds determined potential candidate genes in the thermogenesis-related pathways that were under selection. Specifically, functional analysis identified a substitution of PRDM16 (p.P779L) in northern cattle, which maintains brown adipocyte formation by boosting thermogenesis-related gene expression, indicating a vital role of this gene in cold tolerance. These findings provide a basis for genetic variation in domestic cattle shaped by environmental temperature and highlight the role of reverse mutation in livestock species.

Ancient Demographics Determine the Effectiveness of Genetic Purging in Endangered Lizards.

The purging of deleterious alleles has been hypothesized to mitigate inbreeding depression, but its effectiveness in endangered species remains debatable. To understand how deleterious alleles are purged during population contractions, we analyzed genomes of the endangered Chinese crocodile lizard (Shinisaurus crocodilurus), which is the only surviving species of its family and currently isolated into small populations. Population genomic analyses revealed four genetically distinct conservation units and sharp declines in both effective population size and genetic diversity. By comparing the relative genetic load across populations and conducting genomic simulations, we discovered that seriously deleterious alleles were effectively purged during population contractions in this relict species, although inbreeding generally enhanced the genetic burden. However, despite with the initial purging, our simulations also predicted that seriously deleterious alleles will gradually accumulate under prolonged bottlenecking. Therefore, we emphasize the importance of maintaining a minimum population capacity and increasing the functional genetic diversity in conservation efforts to preserve populations of the crocodile lizard and other endangered species.

Development and Forensic Application of EX20+30Y Kit. / EX20+30Y试剂盒的建立及法医学应用.

OBJECTIVES: To develop a multiplex PCR amplification system (EX20+30Y for short) of 19 autosomes, 30 Y-STR loci plus the gender indicator, and evaluate its forensic application value. METHODS: With the six-color fluorescence labeling technology, a multiplex amplification system of 19 autosomal STR loci and 30 Y-STR loci plus the gender indicator was constructed. Blood samples from 210 unrelated individuals, 69 daily case samples and standard samples 9948 and 9947A were collected for loci detection and analysis. The EX20+30Y multiplex amplification system was evaluated by its sensitivity, mixed sample detection ability, species specificity, balance, direct amplification ability, sample applicability and anti-inhibition ability. RESULTS: Multiplex amplification of blood samples from 210 unrelated individuals by the detection system obtained accurate genotyping results. The detection sensitivity of standard samples was 0.125 ng and the species specificity was high. The 69 samples from daily cases were genotyped correctly. Moreover, standard sample 9948 could be accurately genotyped even if the samples contained a certain concentration of inhibitors. CONCLUSIONS: The multiplex amplification system established in this study can conduct combined examination of 19 autosomes, 30 Y-STR loci plus the gender indicator with accurate genotyping and high sensitivity. It has a good forensic application prospect.

Phosphoribosyl-pyrophosphate synthetase 2 (PRPS2) depletion regulates spermatogenic cell apoptosis and is correlated with hypospermatogenesis.

Phosphoribosyl-pyrophosphate synthetase 2 (PRPS2) is a rate-limiting enzyme and plays an important role in purine and pyrimidine nucleotide synthesis. Recent studies report that PRPS2 is involved in male infertility. However, the role of PRPS2 in hypospermatogenesis is unknown. In this study, the relationship of PRPS2 with hypospermatogenesis and spermatogenic cell apoptosis was investigated. The results showed that PRPS2 depletion increased the number of apoptotic spermatogenic cells in vitro. PRPS2 was downregulated in a mouse model of hypospermatogenesis. When PRPS2 expression was knocked down in mouse testes, hypospermatogenesis and accelerated apoptosis of spermatogenic cells were noted. E2F transcription factor 1 (E2F1) was confirmed as the target gene of PRPS2 and played a key role in cell apoptosis by regulating the P53/Bcl-xl/Bcl-2/Caspase 6/Caspase 9 apoptosis pathway. Therefore, these data indicate that PRPS2 depletion contributes to the apoptosis of spermatogenic cells and is associated with hypospermatogenesis, which may be helpful for the diagnosis of male infertility.

Molecular structure, expression analysis and functional characterization of APRIL (TNFSF 13) in goat (Capra hircus).

A proliferation-inducing ligand (APRIL) is an important member of the tumor necrosis factor (TNF) superfamily. In the present study, a novel cDNA was isolated from the spleen of goat by RT-PCR and designated as goat APRIL (gAPRIL). The open reading frame (ORF) of this cDNA covered 753bp, encoding a protein of 250 amino acids. Sequence comparison showed that gAPRIL contains a predicted transmembrane domain, a putative furin protease cleavage site, and two cysteine residues, which are the typical characteristics of TNF gene in mammals. The predicted three dimensional (3D) structure of soluble part of the gAPRIL (gsAPRIL) monomer analyzed by comparative protein modeling revealed that it is very similar to its counterparts. Real-time PCR analysis revealed that gAPRIL was constitutively expressed in various tissues. Recombinant gsAPRIL fused with NusA tag was efficiently produced in Escherichia coli BL21 (DE3) and then analyzed by the SDS-PAGE as well as western blot. Laser scanning confocal microscopy analysis showed gsAPRIL could bind to its receptors. In vitro, the MTT and flow cytometric methods revealed that purified gsAPRIL protein was not only able to promote survival/proliferation of goat splenocytes, but also able to stimulate survival/proliferation of mouse B cells. These results indicated that gAPRIL plays an important role in survival/proliferation of goat splenocytes and provided a basis for investigating its potential to be used as an immunoadjuvant for enhancing vaccine efficacy and as an immunotherapeutic in goats.

Molecular structure, phylogenetic analysis, tissue distribution, and function characterization of interferon-γ-inducible lysosomal thiol reductase (GILT) gene in sheep (Ovis aries).

Interferon-γ-inducible-lysosomal thiol reductase (GILT) plays a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction. In this study, a sheep cDNA, sGILT, encoding GILT protein was isolated from the spleen cDNA library of Ovis aries. It codes for a deduced protein of 244 amino acids with a putative molecular weight of 27.6 kDa, which has all the typical structural features of GILT protein including an active-site CXXC motif, a GILT signature sequence CQHGX(2)ECX(2)NX(4)C, and 5 conserved cysteines. Sequence comparison indicated the amino acid sequence of sGILT showed high identity to cow GILT (93.03%). Phylogenetic analysis showed that sGILT and cow GILT shared the greatest homology. The result of real-time PCR suggested that sGILT mRNA was expressed in a tissue-specific manner and obviously up-regulated in splenocytes and PBMCs after induction with lipopolysaccharide (LPS). Recombinant sGILT fused with His(6) tag was efficiently expressed in Escherichia coli BL21 (DE3) and purified by Ni-NTA affinity chromatography. Its expression was confirmed by SDS-PAGE and Western blotting analysis. Thiol reductase activity was assessed using antibody as the substrate. These results suggest that sGILT has the activity of disulfide bond reduction and indicate that sheep also express a protein that has been found to maintain first line of innate immune defense at basal level.