Enzyme Reaction-Assisted Programmable Transcriptional Switches for Bioactive Molecule Detection.

Bioactive molecules are highly worthwhile to recognize and explore the latent pathogenic mechanism. Conventional methods for bioactive molecule detection, including mass spectrometry and fluorescent probe imaging, are limited due to the complex processing and signal interference. Here, we designed enzyme-reaction-assisted programmable transcriptional switches for the detection of bioactive molecules. The approach is based on the use of programmable enzyme site-specific cleavage-assisted DNA triplex-based conformational switches that, upon responding to bioactive molecules, can trigger the transcription of fluorescent light-up aptamers. Thanks to the programmable nature of the sensing platform, the method can be adapted to different bioactive molecules, and we demonstrated the enzyme-small molecule catalytic reaction combination of myeloperoxidase (MPO)-hydrogen peroxide (H2O2) as a model that transcriptional switches was capable of detecting H2O2 and possessed the specificity and anti-interference ability in vitro. Furthermore, we successfully applied the switches into cells to observe the detection feasibility in vivo, and dynamically monitored changes of H2O2 in cellular oxidative stress levels. Therefore, we attempt to amalgamate the advantages of enzyme reaction with the pluripotency of programmable transcriptional switches, which can take both fields a step further, which may promote the research of biostimuli and the construction of DNA molecular devices.

Paper-based netlike rolling circle amplification (NRCA) for ultrasensitive and visual detection of SARS-CoV-2.

COVID-19 is a highly diffuse respiratory infection caused by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2). Currently, quantitative real-time polymerase chain reaction (qRT-PCR) technology is commonly used in clinical diagnosis of COVID-19. However, this method is time-consuming and labor-intensive, which is limited in clinical application. Here, we propose a new method for the ultrasensitive and visual detection of SARS-CoV-2 viral nucleic acid. The assay integrates with a paper device and highly efficient isothermal amplification technology - Netlike rolling circle amplification (NRCA), which can reach a limit of detection of 4.12 aM. The paper-based NRCA owns advantages of specificity, portability, visualization and low-cost. Therefore, this method can effectively meet the requirements of point-of-care testing, providing a novel molecular detection technology for clinical diagnosis of COVID-19 and promoting the development of NRCA devices.

Established risk prediction models for the incidence of a low lean tissue index in patients with peritoneal dialysis.

OBJECTIVE: The objective of this study is to investigate the incidence of low lean tissue index (LTI) and the risk factors for low LTI in peritoneal dialysis (PD) patients, including to establish risk prediction models. METHODS: A total of 104 PD patients were enrolled from October 2019 to 2021. LTI was measured by bioimpedance spectroscopy. Multivariate logistic regression and machine learning were used to analyze the risk factors for low LTI in PD patients. Kaplan-Meier analysis was used to analyze the survival rate of patients with low LTI. RESULTS: The interleukin-6 (IL-6) level, red cell distribution width (RDW), overhydration, body mass index (BMI), and the subjective global assessment (SGA) rating significantly differed between the low LTI and normal LTI groups (all p < 0.05). Multivariate logistic regression showed that IL-6 (1.10 [95% CI: 1.02-1.18]), RDW (1.87 [95% CI: 1.18-2.97]), BMI (0.97 [95% CI: 0.68-0.91]), and the SGA rating (6.33 [95% CI: 1.59-25.30]) were independent risk factors for LTI. Cox regression analysis showed that low LTI (HR 3.14, [95% CI: 1.12-8.80]) was the only significant risk factor for all-cause death in peritoneal dialysis patients. The decision process to predict the incidence of low LTI in PD patients was established by machine learning, and the area under the curve of internal validation was 0.6349. CONCLUSIONS: Low LTI is closely related to mortality in PD patients. Microinflammatory status, high RDW, low BMI and low SGA rating are risk factors for low LTI in PD patients. The developed prediction model may serve as a useful tool for assessing low LTI in PD patients.

Genome-wide identification of ZmSnRK2 genes and functional analysis of ZmSnRK2.10 in ABA signaling pathway in maize (Zea mays L).

BACKGROUND: Phytohormone abscisic acid (ABA) is involved in the regulation of a wide range of biological processes. In Arabidopsis, it has been well-known that SnRK2s are the central components of the ABA signaling pathway that control the balance between plant growth and stress response, but the functions of ZmSnRK2 in maize are rarely reported. Therefore, the study of ZmSnRK2 is of great importance to understand the ABA signaling pathways in maize. RESULTS: In this study, 14 ZmSnRK2 genes were identified in the latest version of maize genome database. Phylogenetic analysis revealed that ZmSnRK2s are divided into three subclasses based on their diversity of C-terminal domains. The exon-intron structures, phylogenetic, synteny and collinearity analysis indicated that SnRK2s, especially the subclass III of SnRK2, are evolutionally conserved in maize, rice and Arabidopsis. Subcellular localization showed that ZmSnRK2 proteins are localized in the nucleus and cytoplasm. The RNA-Seq datasets and qRT-PCR analysis showed that ZmSnRK2 genes exhibit spatial and temporal expression patterns during the growth and development of different maize tissues, and the transcript levels of some ZmSnRK2 genes in kernel are significantly induced by ABA and sucrose treatment. In addition, we found that ZmSnRK2.10, which belongs to subclass III, is highly expressed in kernel and activated by ABA. Overexpression of ZmSnRK2.10 partially rescued the ABA-insensitive phenotype of snrk2.2/2.3 double and snrk2.2/2.3/2.6 triple mutants and led to delaying plant flowering in Arabidopsis. CONCLUSION: The SnRK2 gene family exhibits a high evolutionary conservation and has expanded with whole-genome duplication events in plants. The ZmSnRK2s expanded in maize with whole-genome and segmental duplication, not tandem duplication. The expression pattern analysis of ZmSnRK2s in maize offers important information to study their functions. Study of the functions of ZmSnRK.10 in Arabidopsis suggests that the ABA-dependent members of SnRK2s are evolutionarily conserved in plants. Our study elucidated the structure and evolution of SnRK2 genes in plants and provided a basis for the functional study of ZmSnRK2s protein in maize.

[Cytogenetic and molecular analysis of a case of Prader-Willi syndrome].

OBJECTIVE: To investigate the significance of cytogenetic and molecular genetic diagnosis of a special type of secondary sexual dysplasia and the applicability of various methods for its detection. METHODS: Using karyotype analysis, array comparative genomic hybridization (aCGH), multiplex ligation-dependent probe amplification (MLPA) and methylation-specific PCR (MS-PCR), we diagnosed and differentially diagnosed a case of secondary sexual dysplasia. RESULTS: Abnormalities were not found in the karyotype analysis or the SRY and AZF gene detection, nor chromosomal duplication and deletion in the initial SurePrint G3 Human CGH Array Kit8×60K.SurePrint G3 unrestricteda CGH ISCA v2,88×60K, however, identified a 68.9 kb deletion of chromosome 15 (hg19:25190737-25259677). MLPA revealed the deletion of exon 3 of the SNRPN gene. MS-PCR showed a significant decrease in the paternal fragment signals, but no difference in the maternal fragment signals between the sample from the patient and that from the control. CONCLUSIONS: The patient was confirmed with Prader-Willi syndrome by various methods of detection.

[Application of array comparative genomic hybridization in prenatal diagnosis of a case with 5q35 deletion syndrome].

OBJECTIVE: To use combined G-banding and array-comparative genomic hybridization (aCGH) for the prenatal diagnosis of a fetus with 5q35 deletion syndrome. METHODS: Chromosomal karotypes of the fetus and parents were analyzed with G-banding analysis. aCGH was performed to detect minor chromosomal structural abnormalities. RESULTS: The karyotype of the fetus was ascertained as 46, XY, t(5;10)(q35;p13), and the karyotypes of the parents were normal. aCGH has identified a de novo 1.68 Mb deletion at 5q35.2q35.3 and a 1.44 Mb duplication at 10p14p13. CONCLUSION: aCGH has a higher resolution and greater accuracy for mapping chromosomal aberrations and is a useful supplement for G banding karyptyping analysis.

[Association of SPO11 and GST gene polymorphisms with idiopathic male infertility in ethnic Han Chinese].

OBJECTIVE: To explore the possible roles of polymorphisms of SPO11 and glutathionine S-transferase (GST) genes in idiopathic male infertility in a ethnic Han Chinese population from Henan. METHODS: Multiplex PCR and DNA sequencing were performed to determine the SPO11 c.517C>T(rs28368082) and GST genes (GSTM1, GSTT1, GSTP1) polymorphisms in 216 idiopathic male infertility cases and 198 normal samples. RESULTS: The frequencies of the SPO11 CC and CT genotypes were 87.5% (189/216) and 12.5% (27/216) in the patients, and 97.5% (193/198) and 2.5% (5/198) in the controls, respectively. The frequencies of SPO11 CC and CT genotypes, the A>G transition at nucleotide 313 in the exon 5 of the GSTP1 gene, and the frequencies of combined genotypes GSTM1 (-/-), GSTT1 (+/+), GSTP1 (AA) and SPO11 (CT) were significantly different between the two groups (P<0.05). CONCLUSION: The rs28368082 polymorphism of the SPO11 gene, the A>G transition at nucleotide 313 in the exon 5 of the GSTP1 gene, and the combined genotypes of GSTM1 (-/-), GSTT1 (+/+), GSTP1 (AA) and SPO11 (CT) may be associated with idiopathic male infertility in ethnic Han Chinese.

Catalytic hairpin assembly-based AIEgen/graphene oxide nanocomposite for fluorescence-enhanced and high-precision spatiotemporal imaging of microRNA in living cells.

The low abundance, heterogeneous expression, and temporal changes of miRNA in different cellular locations pose significant challenges for both the detection sensitivity of miRNA liquid biopsy and intracellular imaging. In this work, we report an intelligently assembled biosensor based on catalytic hairpin assembly (CHA) and aggregation-induced emission (AIE), named as catalytic hairpin aggregation-induced emission (CHAIE), for the ultrasensitive detection and intracellular imaging of miRNA-155. To achieve such goal, tetraphenylethylene-N3 (TPE-N3) is used as AIE luminogen (AIEgen), while graphene oxide is introduced to quench the fluorescence. When the target miRNA is present, CHA reaction is triggered, causing the AIEgen to self-assemble with the hairpin DNA. This will restrict the intramolecular rotation of the AIEgen and produce a strong AIE fluorescence. Interestingly, CHAIE does not require any enzyme or expensive thermal cycling equipment, and therefore provides a rapid detection. Under optimal conditions, the proposed biosensor can determine miRNA in the concentration range from 2 pM to 200 nM within 30 min, with the detection limit of 0.42 pM. The proposed CHAIE biosensor in this work offers a low background signal and high sensitivity, making it applicable for highly precise spatiotemporal imaging of target miRNA in living cells.

Synthetic biology-based bioreactor and its application in biochemical analysis.

In the past few years, synthetic biologists have established some biological elements and bioreactors composed of nucleotides under the guidance of engineering methods. Following the concept of engineering, the common bioreactor components in recent years are introduced and compared. At present, biosensors based on synthetic biology have been applied to water pollution monitoring, disease diagnosis, epidemiological monitoring, biochemical analysis and other detection fields. In this paper, the biosensor components based on synthetic bioreactors and reporters are reviewed. In addition, the applications of biosensors based on cell system and cell-free system in the detection of heavy metal ions, nucleic acid, antibiotics and other substances are presented. Finally, the bottlenecks faced by biosensors and the direction of optimization are also discussed.

Study on the effect of compound cultivation on the growth feature and active ingredients content of Salvia miltiorrhiza.

We investigated the effects of the complex cultivation of Salvia miltiorrhiza on microbial communities, secretions, yield, and active ingredients, and the mechanism of action between microbial communities, secretions, and S. miltiorrhiza growth and development. Neither maize nor soybean was suitable to grow with S. miltiorrhiza, but sesame significantly increased salvinone content, the active ingredient of S. miltiorrhiza, and Tanshinone IIA, Tanshinone I, and Cryptotanshinone increased by 27.06%, 22.76%, and 26.41%, respectively, which increased the abundance and number of microbial communities in S. miltiorrhiza roots. 16S rRNA results showed that the most abundant bacterial phyla were Proteobacteria and Acidobacteriota, and their number increased with compound planting of sesame and S. miltiorrhiza. Salvia inter-root secretions affected the microbial community and Salvia growth and development, and lipids and lipid-like molecules significantly reduced Salvia yield and active ingredients. Overall, different plant secretions can lead to differences in the natural environment and Salvia root growth and development, and the composite planting of sesame with Salvia can improve inter-root microbial communities, enhance Salvia quality, and make fuller use of land resources.

A peptide-DNA hybrid bio-nanomicelle and its application for detection of caspase-3 activity.

Bio-nanomicelles based on biomaterials such as nucleic acids, peptides, glycans, and lipids have developed rapidly in the field of bioanalysis. Although DNA and peptides have unique advantages, unfortunately, there are few bio-nanomicelles integrating DNA with peptides. Here, we designed a peptide-DNA hybrid bio-nanomicelle for the activity detection of caspase-3. The detection mechanism is based on caspase-3 specific recognition and cleavage of peptide substrates, which owns high sensitivity and selectivity. Under optimal conditions, the detection of caspase-3 activity can be achieved using our designed bio-nanomicelles and the detection limit is 0.72 nM. Furthermore, the proposed method was also successfully applied for the detection of caspase-3 in cell lysate samples after apoptosis-inducing.

Prognostic Value of Pretreatment Prognostic Nutritional Index in Patients With Renal Cell Carcinoma: A Meta-Analysis.

BACKGROUND: The pretreatment prognostic nutritional index (PNI) is correlated with poor prognosis in several malignancies. However, the prognostic role of PNI in patients with renal cell carcinoma (RCC) remains unclear. Therefore, we performed a meta-analysis to investigate the prognostic significance of PNI in patients with RCC. METHODS: We searched the PubMed, Web of Science, Embase, Scopus, and Cochrane Library databases up to February 2021. Pooled hazard ratios (HRs) and 95% confidence intervals (CIs) were used to estimate correlation between PNI and survival endpoints in RCC. RESULTS: Ten studies with 4,908 patients were included in the meta-analysis. The pooled results indicated that a low PNI associated with poor overall survival (HR = 2.10, 95% CI = 1.67-2.64, p<0.001), shorter progression-free survival, disease-free survival, recurrence-free survival (HR = 1.99, 95% CI = 1.67-2.36, p<0.001), and poor cancer-specific survival (HR = 2.95, 95% CI = 1.61-5.39, p<0.001). Additionally, the prognostic ability of PNI was not affected by subgroup analysis factors. CONCLUSION: The meta-analysis indicated that low PNI associated with shorter survival outcomes in patients with RCC. Therefore, PNI could be used as an effective prognostic indicator in RCC.