Comparative analysis of the complete chloroplast genome of Pueraria provides insights for species identification, phylogenetic relationships, and taxonomy.
Pueraria is an edible and medicinal raw material, which is of great value to the pharmaceutical and food industries. Nonetheless, due to morphological diversity and complex domestication history, the classification of Pueraria plants is ambiguous. As the varieties on the market are mixed, the species are difficult to distinguish, and their morphological characteristics are similar to the physical and chemical properties. It is difficult to accurately identify them by traditional identification methods. Chloroplast (cp) genomes are widely used in species identification and phylogenetic studies to achieve accurate identification of medicinal plants, and can also provide more reference information for phylogenetic studies. Based on interspecific and intraspecific sampling, the cp genomes of eight species or varieties of Pueraria plants were examined in this study.
The study unveiled that the cp genome size varied from 151,555 to 153,668 base pairs (bp), with the total GC content ranging from 35.4 to 37.0%. Moreover, it was discerned that the cp genome contained between 128 and 135 genes. Comparative analysis indicated that the highest number of Simple Sequence Repeats (SSRs) was identified in P. montana and P. alopecuroides, with a preponderance of these SSRs being rich in Adenine (A) and Thymine (T) nucleotides. Complete comparison and sliding window analysis of the cp genome established that the non-coding region exhibited greater sequence differences than the coding region, and that the large single copy (LSC) region demonstrated higher nucleotide polymorphism levels. Fourteen highly variable loci such as rpoB,ycf1,rbcL,trnF-GAA-trnL,psbC-psbD, and ycf4-cemA were detected as potential molecular markers for Pueraria species identification. Moreover, the phylogenetic tree demonstrated that other Pueraria species had the most distant relationship with Haymondia wallichii and Toxicopueraria peduncularis, thereby offering fresh perspectives into the species classification of Pueraria. The molecular clock analysis results indicate that the divergence time of Pueraria may occur at ∼6.46 Ma. It is speculated that the cold climate may be the cause of Pueraria species diversity and promote the radiation of the genus.
This research provides theoretical backing and serves as a reference point for the identification and taxonomical classification of Pueraria species. The findings will prove beneficial in future studies on the preservation of medicinal resources, phylogenetic relationships, and genetic engineering of Pueraria plants.
Hai Y
,Huang X
,Sun H
,Sun J
,Li J
,Zhang Y
,Qian Y
,Wu J
,Yang Y
,Xia C
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《BMC PLANT BIOLOGY》
Comparative chloroplast genome analysis of Ardisia (Myrsinoideae, Primulaceae) in China and implications for phylogenetic relationships and adaptive evolution.
Numerous species of Ardisia are widely used for their medicinal and ornamental values in China. However, accurately identifying Ardisia species at the molecular level remains a challenge due to the morphological similarities among different species, the complexity of interspecific variation, and the limited availability of genetic markers. In this study, we reported 20 chloroplast genomes of Ardisia species from China and combined them with 8 previously published chloroplast genomes to conduct a comprehensive analysis for phylogenetic relationships and adaptive evolution.
For the 28 Ardisia species analyzed in this study, the size of the chloroplast genomes ranged from 155,088 bp to 156,999 bp, and all exhibited a typical tetrad structure with conserved gene content and number. Each genome contained 85-88 protein-coding genes, 36-37 tRNA genes, and 8 rRNA genes. Comparative analysis showed that the genomic structures and gene order were relatively conserved with slight variations in the inverted repeat regions (IRs). Simple sequence repeats (SSRs) were predominantly single nucleotide repeats, while repeat sequences were mainly composed of palindromic and forward repeats. Twelve highly variable regions were identified as potential DNA barcodes for species identification and phylogenetic analysis of Ardisia. The phylogenetic tree supported the division of the subgenus Bladhia s.l. into two subgenera: Bladhia s.str. and Odontophylla (Yang) Huang. Further investigation revealed that two protein-coding genes (rbcL and rpoC2) were under positive selection and might be associated with the adaptation of Ardisia species to shaded environments.
Our study analyzed the chloroplast genomes of 20 Ardisia species from China to explore their phylogenetic relationships and adaptive evolution. By combining these results with data from eight previously published chloroplast genomes, the essential characteristics of Ardisia chloroplast genomes were clarified. The research establishes a theoretical basis for the classification, identification, and comprehension of the adaptive evolution of Ardisia species.
Zhang J
,Ning Y
,Li J
,Deng Y
,Wang L
,Mao S
,Zhao B
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《BMC PLANT BIOLOGY》
Can a Liquid Biopsy Detect Circulating Tumor DNA With Low-passage Whole-genome Sequencing in Patients With a Sarcoma? A Pilot Evaluation.
A liquid biopsy is a test that evaluates the status of a disease by analyzing a sample of bodily fluid, most commonly blood. In recent years, there has been progress in the development and clinical application of liquid biopsy methods to identify blood-based, tumor-specific biomarkers for many cancer types. However, the implementation of these technologies to aid in the treatment of patients who have a sarcoma remains behind other fields of cancer medicine. For this study, we chose to evaluate a sarcoma liquid biopsy based on circulating tumor DNA (ctDNA). All human beings have normal cell-free DNA (cfDNA) circulating in the blood. In contrast with cfDNA, ctDNA is genetic material present in the blood stream that is derived from a tumor. ctDNA carries the unique genomic fingerprint of the tumor with changes that are not present in normal circulating cfDNA. A successful ctDNA liquid biopsy must be able to target these tumor-specific genetic alterations. For instance, epidermal growth factor receptor (EGFR) mutations are common in lung cancers, and ctDNA liquid biopsies are currently in clinical use to evaluate the status of disease in patients who have a lung cancer by detecting EGFR mutations in the blood. As opposed to many carcinomas, sarcomas do not have common recurrent mutations that could serve as the foundation to a ctDNA liquid biopsy. However, many sarcomas have structural changes to their chromosomes, including gains and losses of portions or entire chromosomes, known as copy number alterations (CNAs), that could serve as a target for a ctDNA liquid biopsy. Murine double minute 2 (MDM2) amplification in select lipomatous tumors or parosteal osteosarcoma is an example of a CNA due to the presence of extra copies of a segment of the long arm of chromosome 12. Since a majority of sarcomas demonstrate a complex karyotype with numerous CNAs, a blood-based liquid biopsy strategy that searches for these CNAs may be able to detect the presence of sarcoma ctDNA. Whole-genome sequencing (WGS) is a next-generation sequencing technique that evaluates the entire genome. The depth of coverage of WGS refers to how detailed the sequencing is, like higher versus lower power on a microscope. WGS can be performed with high-depth sequencing (that is, > 60×), which can detect individual point mutations, or low-depth sequencing (that is, 0.1× to 5×), referred to as low-passage whole-genome sequencing (LP-WGS), which may not detect individual mutations but can detect structural chromosomal changes including gains and losses (that is, CNAs). While similar strategies have shown favorable early results for specific sarcoma subtypes, LP-WGS has not been evaluated for applicability to the broader population of patients who have a sarcoma.
Does an LP-WGS liquid biopsy evaluating for CNAs detect ctDNA in plasma samples from patients who have sarcomas representing a variety of histologic subtypes?
This was a retrospective study conducted at a community-based, tertiary referral center. Nine paired (plasma and formalin-fixed paraffin-embedded [FFPE] tissue) and four unpaired (plasma) specimens from patients who had a sarcoma were obtained from a commercial biospecimen bank. Three control specimens from individuals who did not have cancer were also obtained. The paired and unpaired specimens from patients who had a sarcoma represented a variety of sarcoma histologic subtypes. cfDNA was extracted, amplified, and quantified. Libraries were prepared, and LP-WGS was performed using a NextSeq 500 next-generation sequencing machine at a low depth of sequencing coverage (∼1×). The ichorCNA bioinformatics algorithm, which was designed to detect CNAs from low-depth genomic sequencing data, was used to analyze the data. In contrast with the gold standard for diagnosis in the form of histopathologic analysis of a tissue sample, this test does not discriminate between sarcoma subtypes but detects the presence of tumor-derived CNAs within the ctDNA in the blood that should not be present in a patient who does not have cancer. The liquid biopsy was positive for the detection of cancer if the ichorCNA algorithm detected the presence of ctDNA. The algorithm was also used to quantitatively estimate the percent ctDNA within the cfDNA. The concentration of ctDNA was then calculated from the percent ctDNA relative to the total concentration of cfDNA. The CNAs of the paired FFPE tissue and plasma samples were graphically visualized using aCNViewer software.
This LP-WGS liquid biopsy detected ctDNA in 9 of 13 of the plasma specimens from patients with a sarcoma. The other four samples from patients with a sarcoma and all serum specimens from patients without cancer had no detectable ctDNA. Of those 9 patients with positive liquid biopsy results, the percent ctDNA ranged from 6% to 11%, and calculated ctDNA quantities were 0.04 to 5.6 ng/mL, which are levels to be expected when ctDNA is detectable.
In this small pilot study, we were able to detect sarcoma ctDNA with an LP-WGS liquid biopsy searching for CNAs in the plasma of most patients who had a sarcoma representing a variety of histologic subtypes.
These results suggest that an LP-WGS liquid biopsy evaluating for CNAs to identify ctDNA may be more broadly applicable to the population of patients who have a sarcoma than previously reported in studies focusing on specific subtypes. Large prospective clinical trials that gather samples at multiple time points during the process of diagnosis, treatment, and surveillance will be needed to further assess whether this technique can be clinically useful. At our institution, we are in the process of developing a large prospective clinical trial for this purpose.
Anderson CJ
,Yang H
,Parsons J
,Ahrens WA
,Jagosky MH
,Hsu JH
,Patt JC
,Kneisl JS
,Steuerwald NM
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