Sequenced-based Rwanda population provides insights into demographic history.

Rwanda is known as the heart of Africa, reflecting the history of the world. Colonization and genocide have led to Rwanda's existing genetic structure. Herein, we used massively parallel sequencing to analyze 296 loci in 185 Rwandans and constructed a database for Rwandan forensic data for the first time. We found the following results: First, forensic parameters demonstrated that all loci were highly informative and could be used for forensic identification and paternity tests in Rwandans. Second, we found that the differences in genetic background between Rwandans and other African populations were similar but slight, as indicated by the massively parallel sequencing panel. Rwandans belonged to the African population and were inseparable from populations from neighboring countries. Also, Rwandans were closer to the European and American populations because of colonization, war, and other reasons. There was no scientific basis for racial classification established by colonization. Further research still needs to be carried out on more loci and larger Rwandan samples.

A novel forensic panel of 186-plex SNPs and 123-plex STR loci based on massively parallel sequencing.

The MiSeq® FGX Forensic system and the HID-Ion AmpliSeq Panel were previously developed for massively parallel sequencing (MPS) for forensic casework. Among the three major sequencing platforms, BGISEQ-500TM, which is based on multiple PCRs, is still lacking in forensics. Here, a novel forensic panel was constructed to detect 186 single-nucleotide polymorphisms (SNPs) and 123 short tandem repeats (STRs) with MPS technology on the BGISEQ-500™ platform. First, the library preparation, sequencing process, and data analysis were performed, focusing on the average depth of coverage and heterozygote balance. We calculated the allelic frequencies and forensic parameters of STR and SNP loci in 73 unrelated Chinese Han individuals. In addition, performance was evaluated with accuracy, uniformity, sensitivity, PCR inhibitor, repeatability and reproducibility, mixtures, degraded samples, case-type samples, and pedigree analyses. The results showed that 100% accurate and concordant genotypes can be obtained, and the loci with an abundance in the interquartile range accounted for 92.90% of the total, suggesting reliable uniformity in this panel. We obtained a locus detection rate that was higher than 98.78% from 78 pg of input DNA, and the optimal amount was 1.25-10 ng. The maximum concentrations of hematin and humic acid were 200 and 100 µM, respectively (the ratios of detected loci were 96.52% and 92.41%), in this panel. As a mixture, compared with those of SNPs, minor-contributor alleles of STRs could be detected at higher levels. For the degraded sample, the ratio of detected loci was 98.41%, and most profiles from case-type samples were not significantly different in abundance in our studies. As a whole, this panel showed high-performance, reliable, robust, repeatable, and reproducible results, which are sufficient for paternity testing, individual identification, and use for potentially degraded samples in forensic science.

Development and Verification of an Economical Method of Custom Target Library Construction.

Although technological advances have greatly reduced the cost of DNA sequencing, sample preparation time and reagent costs remain the limiting factors for many studies. Based on low-cost targeted amplification, we developed an economical method for custom target library construction based on DNA nanoball (DNB) technology and two-step polymerase chain reaction (PCR). Here, we refer to this method as the two-step PCR, which was compared to traditional multiplex PCR methods in three aspects, data quality, efficiency, and specificity to humans. The results confirmed that two-step PCR reduces to finishing 128 sequencing libraries in only 2 h 24 min 59 s of the total PCR time and at a data utilization rate of 0.44 at a cost of approximately $1.70 per sample for targeted sequencing via the two-step PCR. The replacement of traditional multiplex PCR methods with this strategy makes the sample preparation process before sequencing relatively more cost-effective and further reduces the cost of next-generation sequencing (NGS). This method may also be free from the interference of other species and the limitations of sample type and DNA content. These findings reveal possibilities for broad applications of this approach in forensic research.

Study of the pharmacokinetics of polymerized porcine hemoglobin (pPolyHb).

Polymerized porcine hemoglobin (pPolyHb), a kind of glutaraldehyde-polymerized haemoglobin-based oxygen carrier, was developed as a potential red blood substitute for clinical applications. Assessment of its absorption, distribution, and metabolism is a major determinant of its safety and efficacy. In this study, a series of pharmacokinetic parameters of pPolyHb were measured and calculated in different animal models, such as the top-load model, the 50% exchange transfusion model and the haemorrhagic shock model. The results showed that the kinetic process of pPolyHb in rats conforms to the laws of linear pharmacokinetics in vivo. The half-life of pPolyHb was superior and more stable under non-pathological conditions, but in clinical situations such as trauma and anaemia, the half-life of pPolyHb may decline. The study of clearance (CL) and the apparent volume of distribution (Vd) of pPolyHb in these three different animal models demonstrated longer residence and a predominant, targeted role of pPolyHb with little accumulation in tissues. Through this study, the superior pharmacokinetic characteristics of pPolyHb have been proved and will aid in the determination of a reasonable dosing regimen and administration interval in clinical situations.

Study of the effects of polymerized porcine hemoglobin (pPolyHb) in an acute anemia rat model.

The objective of this study was to investigate the recovery ability of polymerized porcine hemoglobin (pPolyHb) in a rat model of acute anemia caused by normovolemic hemodilution (ANH). After the ANH procedure, rats were infused with either pPolyHb or red blood cells. The results showed pPolyHb could carry a sufficient amount of oxygen to the tissues to maintain normal aerobic metabolism and hemodynamic stability, without any significant toxic effects on renal and liver function according to pathological, and biochemical analysis. The data suggest pPolyHb may be a good candidate for the treatment of acute anemia in future clinical trials.