Humans ; Class I Phosphatidylinositol 3-Kinases/genetics* ; Breast Neoplasms/diagnosis* ; Mutation ; Female ; China ; DNA Mutational Analysis/methods* ; Genetic Testing/standards*

Humans ; Consensus ; High-Throughput Nucleotide Sequencing/standards* ; Neoplasms/diagnosis* ; Quality Control ; Sequence Analysis, DNA/methods* ; Sequence Analysis, RNA/methods*

Descurainiae Semen Lepidii Semen, also known as Tinglizi, originates from Brassicaceae plants Descurainia sophia or Lepidium apetalum. The former is commonly referred to as "Southern Tinglizi(Descurainiae Semen)", while the latter is known as "Northern Tinglizi(Lepidii Semen)". To scientifically and accurately identify the origin of Tinglizi medicinal materials and traditional Chinese medicine products, this study developed a specific DNA mini-barcode based on chloroplast genome sequences. By combining the DNA mini-barcode with DNA metabarcoding technology, a method for the qualitative and quantitative identification of Tinglizi medicinal materials and Chinese patent medicines was established. In this study, chloroplast genomes of Southern Tinglizi and Northern Tinglizi and seven commonly encountered counterfeit products were downloaded from the GenBank database. Suitable polymorphic regions were identified to differentiate these species, enabling the development of the DNA mini-barcode. Using DNA metabarcoding technology, medicinal material mixtures of Southern and Northern Tinglizi, as well as the most common counterfeit product, Capsella bursa-pastoris seeds, were analyzed to validate the qualitative and quantitative capabilities of the mini-barcode and determine its minimum detection limit. Additionally, the mini-barcode was applied to Chinese patent medicines containing Tinglizi to authenticate their botanical origin. The results showed that the developed mini-barcode(psbB) exhibited high accuracy and specificity, effectively distinguishing between the two authentic origins of Tinglizi and commonly encountered counterfeit products. The analysis of mixtures demonstrated that the mini-barcode had excellent qualitative and quantitative capabilities, accurately identifying the composition of Chinese medicinal materials in mixed samples with varying proportions. Furthermore, the analysis of Chinese patent medicines revealed the presence of the adulterant species(Capsella bursa-pastoris) in addition to the authentic species(Southern and Northern Tinglizi), indicating the occurrence of adulteration in commercially available Tinglizi-containing products. This study developed a method for the qualitative and quantitative identification of multi-origin Chinese medicinal materials and related products, providing a model for research on other multi-origin Chinese medicinal materials.
DNA Barcoding, Taxonomic/methods* ; Drugs, Chinese Herbal/chemistry* ; Drug Contamination ; Genome, Chloroplast ; Medicine, Chinese Traditional

OBJECTIVE: To distinguish the ambiguous genotyping results of human leukocyte antigen (HLA), identify a novel HLA-B allele and analyze the nucleotide sequence. METHODS: A total of 2 076 umbilical core blood samples from the Zhejiang Cord Blood Bank in 2022 were detected using the next generation sequencing technology (NGS) based on the Ion Torrent S5 platform. Among these a rare HLA-B allele with ambiguous combination result containing a base mutation was identified, and was further confimed by the third-generation sequencing (TGS) based on the nanopore technology. RESULTS: The NGS typing result of HLA-B locus showed HLA-B* 46:18, 54:06 or HLA-B*46:01, 54:XX (including a base mutation), and nanopore sequencing confirmed the typing as HLA-B*46:01, 54:XX (including a base mutation). Compared with HLA-B*54:01:01:01, the HLA-B*54:XX allele showed one single nucleotide substitution at position 1014 T>C in exon 6, with no amino acid change. The nucleotide sequence of the novel HLA-B*54:XX has been submitted to the GenBank nucleotide sequence database and the accession number OP853532 was assigned. CONCLUSION A ambiguous genotyping of the HLA-B Locus detected by NGS was distinguished by nanopore sequencing and a new HLA-B allele was successfully identified, which was officially named as HLA-B*54:01:11 by the World Health Organization Nomenclature Committee for Factors of the HLA System.
Humans ; High-Throughput Nucleotide Sequencing ; Alleles ; HLA-B Antigens/genetics* ; Genotype ; Mutation ; Sequence Analysis, DNA ; Base Sequence

OBJECTIVE: To assess the diagnostic efficiency of long-read sequencing (LRS) for the determination of CYP21A1P/CYP21A2 and TNXA/TNXB fusion genotypes among children with 21-hydroxylase deficiency (21-OHD) and explore their clinical characteristics. METHODS: LRS sequencing was carried out on 30 children diagnosed with 21-OHD at the Department of Endocrinology, Fujian Children's Hospital between November 2022 and September 2023 by clinical symptoms or conventional Sanger sequencing combined with multiple ligation-dependent probe amplification (MLPA). The results of the two methods were compared. Clinical data of the children were collected and analyzed. This study has been approved by the Medical Ethics Committee of the Fujian Children's Hospital (Ethic No. 2022ETKLR10024). RESULTS: Of the 30 children with 21-OHD, 11 (36.7%) were found to carry CYP21A1P/CYP21A2 and TNXA/TNXB fusion genes by LRS. The most common type of fused CYP21A1P/CYP21A2 gene was CH-1 (72.7%), and 1 (3.3%) was found to harbor TNXA/TNXB CH-1. Eleven cases (36.7%) were found to carry large deletions by Sanger sequencing combined with MLPA, with the most common one being CYP21A2 exons 1-3 del (72.7%), which was followed by CYP21A2 exons 1-7 del (18.2%). Follow up of 11 patients carrying a fusion gene revealed that 6 were sale wasting (SW) types, 5 were simple virilizing (SV) types, whilst no non-classical (NC) type was found. Four girls had presented with central precocious puberty (CPP). One child carrying TNXA/TNXB CH-1 had presented with CAH-X syndrome. CONCLUSION Compared with Sanger sequencing combined with MLPA detection method, LRS sequencing was able to differentiate the subtypes of CYP21A1P/CYP21A2 and TNXA/TNXB fusion genes, pinpoint the breakpoints of the deletions, and directly determine the cis-trans position without the need to analyze the genotype of the pedigree members, which has provided a reliable method for the typing of 21-OHD. As some fusion genes may retain 21-hydroxylase activity, female carriers may have a higher incidence of CPP.
Humans ; Steroid 21-Hydroxylase/genetics* ; Adrenal Hyperplasia, Congenital/genetics* ; Child ; Female ; Male ; Child, Preschool ; Tenascin/genetics* ; Infant ; Genotype ; Sequence Analysis, DNA/methods* ; Pseudogenes

Artemisiae Scoporiae Herba is derived from Artemisia scoparia or A. capillaris. The accurate identification of the herbs, particularly when dealing with bulk samples, is critical for ensuring the quality and efficacy of the medicinal product. This study aimed to establish a comprehensive molecular approach by combining multiple markers for the precise identification of Artemisiae Scoporiae Herba. The ITS2 from A. scoparia, A. capillaris, and other common Artemisia species were retrieved from GenBank. MEGA was used to build a phylogenetic tree with these sequences, and the effectiveness of ITS2 in species identification was assessed. The analysis revealed that while ITS2 could distinguish Artemisiae Scoporiae Herba from other closely related species of Artemisia, it was insufficient to differentiate between A. scoparia and A. capillaris. To address this limitation, the chloroplast genome of A. capillaris was assembled and compared with the published chloroplast genomes of A. scoparia and A. capillaris, on the basis of which a DNA mini-barcode was developed. The rpoA-rps11 region was selected as the target for the development of mini-barcode due to its potential for distinguishing between these two species. Specific primers were designed to differentiate A. scoparia from A. capillaris. The ITS2 sequences and the newly developed mini-barcode were used together for Sanger sequencing to identify individual samples of Artemisiae Scoporiae Herba, while DNA metabarcoding was employed for the identification of bulk samples. The identification results of representative individual samples and bulk samples from different regions consistently confirmed A. capillaris. This study established a method that combined ITS2 and mini-barcode to identify bulk samples of Artemisiae Scoporiae Herba from different regions. This approach overcomes the limitations of morphological and chemical methods, enhancing species identification accuracy and supporting a stable supply of medicinal materials.
Artemisia/classification* ; DNA Barcoding, Taxonomic/methods* ; Phylogeny ; DNA, Plant/genetics* ; DNA, Ribosomal Spacer/genetics*

Humans ; Consanguinity ; Exome Sequencing ; Mutation/genetics* ; Sequence Analysis, DNA ; Ciliary Motility Disorders ; Axonemal Dyneins/genetics*

Eleutherococcus senticosus is one of the Dao-di herbs in northeast China. In this study, the chloroplast genomes of three E. senticosus samples from different genuine producing areas were sequenced and then used for the screening of specific DNA barcodes. The germplasm resources and genetic diversity of E. senticosus were analyzed basing on the specific DNA barcodes. The chloroplast genomes of E. senticosus from different genuine producing areas showed the total length of 156 779-156 781 bp and a typical tetrad structure. Each of the chloroplast genomes carried 132 genes, including 87 protein-coding genes, 37 tRNAs, and 8 rRNAs. The chloroplast genomes were relatively conserved. Sequence analysis of the three chloroplast genomes indicated that atpI, ndhA, ycf1, atpB-rbcL, ndhF-rpl32, petA-psbJ, psbM-psbD, and rps16-psbK can be used as specific DNA barcodes of E. senticosus. In this study, we selected atpI and atpB-rbcL which were 700-800 bp and easy to be amplified for the identification of 184 E. senticosus samples from 13 genuine producing areas. The results demonstrated that 9 and 10 genotypes were identified based on atpI and atpB-rbcL sequences, respectively. Furthermore, the two barcodes identified 23 genotypes which were named H1-H23. The haplotype with the highest proportion and widest distribution was H10, followed by H2. The haplotype diversity and nucleotide diversity were 0.94 and 1.82×10~(-3), respectively, suggesting the high genetic diversity of E. senticosus. The results of the median-joining network analysis showed that the 23 genotypes could be classified into 4 categories. H2 was the oldest haplotype, and it served as the center of the network characterized by starlike radiation, which suggested that population expansion of E. senticosus occurred in the genuine producing areas. This study lays a foundation for the research on the genetic quality and chloroplast genetic engineering of E. senticosus and further research on the genetic mechanism of its population, providing new ideas for studying the genetic evolution of E. senticosus.
DNA Barcoding, Taxonomic ; Eleutherococcus/genetics* ; Base Sequence ; Chloroplasts/genetics* ; Genetic Variation ; Phylogeny

Enamel formation is a series of complex physiological processes, which are regulated by critical genes spatially and temporally. These processes involve multiple developmental stages covering ages and are prone to suffer signal interference or gene mutations, ultimately leading to developmental defects of enamel (DDE). Epigenetic modifications have important regulatory roles in gene expression during enarnel development. New technologies including high-throughput sequencing, chromatin immunoprecipitation sequencing (ChIP-seq), and DNA methylation chip are emerging in recent years, making it possible to establish genome-wide epigenetic modification profiles during developmental processes. The regulatory role of epigenetic modification with spatio-temporal pattern, such as DNA methylation, histone modification and non-coding RNA, has significantly expanded our understanding of the regulatory network of enamel formation, providing a new theoretical basis of clinical management and intervention strategy for DDE. The present review briefly describes the enamel formation process of human beings' teeth as well as rodent incisors and summarizes the dynamic characteristics of epigenetic modification during enamel formation. The functions of epigenetic modification in enamel formation and DDE are also emphatically discussed.
Humans ; Epigenesis, Genetic ; Developmental Defects of Enamel ; DNA Methylation ; Oligonucleotide Array Sequence Analysis ; Dental Enamel

OBJECTIVE: To analyze the serological characteristics and molecular mechanism for an individual with p phenotype. METHODS: An individual with p phenotype upon blood group identification at Jiaxing Blood Center in May 2021 was analyzed. ABO, RhD and P1PK blood groups and irregular antibodies in her serum were identified using conventional serological methods. The encoding region of α1, 4-galactosyltransferase gene (A4GALT) encoding P1 and Pk antigens was analyzed by polymerase chain reaction-sequence-based typing (PCR-SBT). RESULTS: The individual was A group, RhD positive and had a p phenotype of the P1PK blood group system. Anti-PP1Pk was discovered in her serum. Sequencing analysis revealed that she has harbored a homozygous c.343A>T variant of the A4GALT gene. CONCLUSION The homozygous c.343A>T variant of the A4GALT gene probably underlay the p phenotype in this individual.
Female ; Animals ; Blood Group Antigens ; Homozygote ; Phenotype ; Polymerase Chain Reaction ; Sequence Analysis, DNA