Complete chloroplast genomes of four Physalis species (Solanaceae): lights into genome structure, comparative analysis, and phylogenetic relationships.

BACKGROUND: Physalis L. is a genus of herbaceous plants of the family Solanaceae, which has important medicinal, edible, and ornamental values. The morphological characteristics of Physalis species are similar, and it is difficult to rapidly and accurately distinguish them based only on morphological characteristics. At present, the species classification and phylogeny of Physalis are still controversial. In this study, the complete chloroplast (cp) genomes of four Physalis species (Physalis angulata, P. alkekengi var. franchetii, P. minima and P. pubescens) were sequenced, and the first comprehensive cp genome analysis of Physalis was performed, which included the previously published cp genome sequence of Physalis peruviana. RESULTS: The Physalis cp genomes exhibited typical quadripartite and circular structures, and were relatively conserved in their structure and gene synteny. However, the Physalis cp genomes showed obvious variations at four regional boundaries, especially those of the inverted repeat and the large single-copy regions. The cp genomes' lengths ranged from 156,578 bp to 157,007 bp. A total of 114 different genes, 80 protein-coding genes, 30 tRNA genes, and 4 rRNA genes, were observed in four new sequenced Physalis cp genomes. Differences in repeat sequences and simple sequence repeats were detected among the Physalis cp genomes. Phylogenetic relationships among 36 species of 11 genera of Solanaceae based on their cp genomes placed Physalis in the middle and upper part of the phylogenetic tree, with a monophyletic evolution having a 100% bootstrap value. CONCLUSION: Our results enrich the data on the cp genomes of the genus Physalis. The availability of these cp genomes will provide abundant information for further species identification, increase the taxonomic and phylogenetic resolution of Physalis, and assist in the investigation and utilization of Physalis plants.

Development of Species-Specific SCAR Markers, Based on a SCoT Analysis, to Authenticate Physalis (Solanaceae) Species.

Physalis is an important genus in the Solanaceae family. It includes many species of significant medicinal value, edible value, and ornamental value. However, many Physalis species are easily confused because of their similar morphological traits, which hinder the utilization and protection of Physalis resources. Therefore, it is necessary to create fast, sensitive, and reliable methods for the Physalis species authentication. Intended for that, in this study, species-specific sequence-characterized amplified region (SCAR) markers were developed for accurate identification of the closely related Physalis species P. angulata, P. minima, P. pubescens, and P. alkekengi var. franchetii, based on a simple and novel marker system, start codon targeted (SCoT) marker. A total of 34 selected SCoT primers yielded 289 reliable SCoT loci, of which 265 were polymorphic. Four species-specific SCoT fragments (SCoT3-1404, SCoT3-1589, SCoT5-550, and SCoT36-520) from Physalis species were successfully identified, cloned, and sequenced. Based on these selected specific DNA fragments, four SCAR primers pairs were developed and named ST3KZ, ST3MSJ, ST5SJ, and ST36XSJ. PCR analysis of each of these primer pairs clearly demonstrated a specific amplified band in all samples of the target Physalis species, but no amplification was observed in other Physalis species. Therefore, the species-specific SCAR primer pairs developed in this study could be used as powerful tools that can rapidly, effectively, and reliably identify and differentiate Physalis species.

[Application of DNA molecular marker technologies in study of medicinal Physalis species].

As traditional Chinese medicinal herbs, Physalis plants have a variety of pharmacological activities, such as anti-inflammatory, anti-oxidant, and anti-cancer effects, and have been used for the treatment of malaria, rheumatism, hepatitis, asthma, and cancer. In addition to the medicinal value, many Physalis species are also the high-grade nutrition health care fruits, can be made canned and candied etc. In the study, the application progress of DNA molecular marker technologies in medicinal Physalis plants in recent years was reviewed, in order to provide an important molecular technical basis for the identification, classification and rational development and protection of medicinal Physalis resources.

Application of the Ribosomal DNA ITS2 Region of Physalis (Solanaceae): DNA Barcoding and Phylogenetic Study.

Recently, commercial interest in Physalis species has grown worldwide due to their high nutritional value, edible fruit, and potential medicinal properties. However, many Physalis species have similar shapes and are easily confused, and consequently the phylogenetic relationships between Physalis species are poorly understood. This hinders their safe utilization and genetic resource conservation. In this study, the nuclear ribosomal ITS2 region was used to identify species and phylogenetically examine Physalis. Eighty-six ITS2 regions from 45 Physalis species were analyzed. The ITS2 sequences were aligned using Clustal W and genetic distances were calculated using MEGA V6.0. The results showed that ITS2 regions have significant intra- and inter-specific divergences, obvious barcoding gaps, and higher species discrimination rates (82.2% for both the BLASTA1 and nearest distance methods). In addition, the secondary structure of ITS2 provided another way to differentiate species. Cluster analysis based on ITS2 regions largely concurred with the relationships among Physalis species established by many previous molecular analyses, and showed that most sections of Physalis appear to be polyphyletic. Our results demonstrated that ITS2 can be used as an efficient and powerful marker in the identification and phylogenetic study of Physalis species. The technique provides a scientific basis for the conservation of Physalis plants and for utilization of resources.

Molecular Identification of Dendrobium Species (Orchidaceae) Based on the DNA Barcode ITS2 Region and Its Application for Phylogenetic Study.

The over-collection and habitat destruction of natural Dendrobium populations for their commercial medicinal value has led to these plants being under severe threat of extinction. In addition, many Dendrobium plants are similarly shaped and easily confused during the absence of flowering stages. In the present study, we examined the application of the ITS2 region in barcoding and phylogenetic analyses of Dendrobium species (Orchidaceae). For barcoding, ITS2 regions of 43 samples in Dendrobium were amplified. In combination with sequences from GenBank, the sequences were aligned using Clustal W and genetic distances were computed using MEGA V5.1. The success rate of PCR amplification and sequencing was 100%. There was a significant divergence between the inter- and intra-specific genetic distances of ITS2 regions, while the presence of a barcoding gap was obvious. Based on the BLAST1, nearest distance and TaxonGAP methods, our results showed that the ITS2 regions could successfully identify the species of most Dendrobium samples examined; Second, we used ITS2 as a DNA marker to infer phylogenetic relationships of 64 Dendrobium species. The results showed that cluster analysis using the ITS2 region mainly supported the relationship between the species of Dendrobium established by traditional morphological methods and many previous molecular analyses. To sum up, the ITS2 region can not only be used as an efficient barcode to identify Dendrobium species, but also has the potential to contribute to the phylogenetic analysis of the genus Dendrobium.

Deep sequencing identifies tissue-specific microRNAs and their target genes involving in the biosynthesis of tanshinones in Salvia miltiorrhiza.

Salvia miltiorrhiza is one of the most popular traditional medicinal herbs in Asian nations. Its dried root contains a number of tanshinones, protocatechuic aldehyde, salvianolic acid B and rosmarinic, and is used for the treatment of various diseases. The finding of microRNAs (miRNAs) and their target genes will help understand their biological role on the biosynthesis of tanshinones in S. miltiorrhiza. In the present study, a total of 452 known miRNAs corresponding to 589 precursor miRNAs (pre-miRNAs), and 40 novel miRNAs corresponding to 24 pre-miRNAs were identified in different tissues of S. miltiorrhiza by high-throughput sequencing, respectively. Among them, 62 miRNAs express only in root, 95 miRNAs express only in stem, 19 miRNAs express only in leaf, and 71 miRNAs express only in flower, respectively. By the degradome analysis, 69 targets potentially cleaved by 25 miRNAs were identified. Among them, acetyl-CoA C-acetyltransferase was cleaved by miR5072, and involved in the biosynthesis of tanshinones. This study provided valuable information for understanding the tissue-specific expression patterns of miRNAs in S. miltiorrhiza, and offered a foundation for future studies of the miRNA-mediated biosynthesis of tanshinones.

High-throughput sequencing and degradome analysis identify miRNAs and their targets involved in fruit senescence of Fragaria ananassa.

In non-climacteric fruits, the respiratory increase is absent and no phytohormone is appearing to be critical for their ripening process. They must remain on the parent plant to enable full ripening and be picked at or near the fully ripe stage to obtain the best eating quality. However, huge losses often occur for their quick post-harvest senescence. To understanding the complex mechanism of non-climacteric fruits post-harvest senescence, we constructed two small RNA libraries and one degradome from strawberry fruit stored at 20°C for 0 and 24 h. A total of 88 known and 1224 new candidate miRNAs, and 103 targets cleaved by 19 known miRNAs families and 55 new candidatemiRNAs were obtained. These targets were associated with development, metabolism, defense response, signaling transduction and transcriptional regulation. Among them, 14 targets, including NAC transcription factor, Auxin response factors (ARF) and Myb transcription factors, cleaved by 6 known miRNA families and 6 predicted candidates, were found to be involved in regulating fruit senescence. The present study provided valuable information for understanding the quick senescence of strawberry fruit, and offered a foundation for studying the miRNA-mediated senescence of non-climacteric fruits.

Functional characterization of AtHsp90.3 in Saccharomyces cerevisiae and Arabidopsis thaliana under heat stress.

The function of cytosolic AtHsp90.3 was characterized by complementing the Saccharomyces cerevisiae endogenous Hsp90 genes and overexpressing it in Arabidopsis thaliana. Though AtHsp90.3 supported the yeast growth under heat stress, in Arabidopsis, compared to the wild type, the transgenic plants overexpressing cytosolic AtHsp90.3 were more sensitive to heat stress with a lower germination rate and higher mortality but and more tolerant to high Ca(2+). Transcriptional expression of heat stress transcription factors, AtHsfA1d, AtHsfA7a and AtHsfB1, and two Hsps, AtHsp101 and AtHsp17, was delayed by constitutive overexpression of cytosolic AtHsp90.3 under heat stress. These results indicate that overexpressing AtHsp90.3 impaired plant tolerance to heat stress and proper homeostasis of Hsp90 was critical for cellular stress response and/or tolerance in plants.