Journal: Advanced Science

Article Title: Early Diagnosis of Colorectal Cancer Based on Bisulfite‐free Site‐specific Methylation Identification PCR Strategy: High‐Sensitivity, Accuracy, and Primary Medical Accessibility

doi: 10.1002/advs.202401137

Figure Lengend Snippet: The scheme of early detection of CRC based on CpG sites detected by the ColoC‐mSTEM. a) STEM‐PCR method: GlaI specifically recognizes and cleaves the 5′‐R (5mC) GY‐3′/3′‐YG (5mC) R‐5′ DNA sequence, thus producing a specific 5′ terminal sequence (P1). A tailored‐designed foldable primer (TFP) that hybridizes with P1 and stops at the 5′ end resulting in the TFP extension products (P2) having the FRc sequence at the 3′end. Consequently, the FRc sequence in P2 spontaneously pairs with the FR sequence to generate a construct that can self‐fold (P3) and self‐prime into a complete hairpin structure (P4) without a 3′ end overhang. P4 can initiate PCR amplification, thus enabling the identification of the methylation status of a specific site. The lowercase letter c indicates the antisense of the sequence. b) Assay development. Bioinformatics methods were used to mine CRC‐related methylation sites in the SDC2 and SFRP2 genes in the TCGA database. Assays for identifying target methylation sites were developed based on the principle of STEM‐PCR technology, and the tolerance of the reaction system to clinical sample testing was verified. The sensitivity, specificity, accuracy, and precision of the assays were subsequently verified to ensure that they could be used for subsequent clinical sample testing. c) Validation of the early detection of ColoC‐mSTEM. Four hundred twenty‐nine of the enrolled samples qualified for further analysis. Group 1 was used for preliminary screening of CRC‐related methylation sites in stool DNA through cluster analysis and receiver operating characteristic (ROC) analysis. The methylation sites selected from Group 1 were detected in Group 2 samples to further confirm the clinical diagnostic performance and explore the early diagnostic ability of the identified sites in stool DNA. Finally, logistic regression analysis was conducted to establish a diagnostic model for CRC via the ColoC‐mSTEM, and the clinical diagnostic performance of the methylation sites and traditional detection indicators was compared.

Article Snippet: As expected, according to our multitarget analysis, the SDC2 gene or SDC2‐6 methylation site was always the most common factor due to the good clinical diagnostic performance of SDC2 gene methylation, especially at the SDC2‐6 methylation site.

Techniques: Biomarker Discovery, Sequencing, Construct, Amplification, Methylation, Diagnostic Assay

Journal: Advanced Science

Article Title: Early Diagnosis of Colorectal Cancer Based on Bisulfite‐free Site‐specific Methylation Identification PCR Strategy: High‐Sensitivity, Accuracy, and Primary Medical Accessibility

doi: 10.1002/advs.202401137

Figure Lengend Snippet: Locations selected for developing ColoC‐mSTEM assay. a) The accuracy of the selected loci for distinguishing CRC tissue from normal tissue (top) (1. SDC2‐4; 2. SDC2‐2; 3. SDC2‐3; 4. SFRP2‐6; 5. SFRP2‐1; 6. SFRP2‐2; 7. SFRP2‐4); For Ft7 model, a ROC curve was plotted for CRC versus normal tissues (below). b) The final selected locations for the ColoC‐mSTEM assay.

Article Snippet: As expected, according to our multitarget analysis, the SDC2 gene or SDC2‐6 methylation site was always the most common factor due to the good clinical diagnostic performance of SDC2 gene methylation, especially at the SDC2‐6 methylation site.

Techniques:

Journal: Advanced Science

Article Title: Early Diagnosis of Colorectal Cancer Based on Bisulfite‐free Site‐specific Methylation Identification PCR Strategy: High‐Sensitivity, Accuracy, and Primary Medical Accessibility

doi: 10.1002/advs.202401137

Figure Lengend Snippet: The process of establishing the ColoC‐mSTEM assay. a) TFP design for SDC2‐6. b) The effect of FR and CRc on the amplification efficiency of the positive template P120:120 copies of methylated DNA per test. c) The effect of FR and CRc on the specificity of negative template amplification; N‐100 ng: 100 ng of undigested DNA per test. d) Effect of methoxy modification on amplification performance; P50: 50 copies of methylated DNA per test; N‐30 ng: 30 ng of undigested DNA per test. e) Effect of TFP primer concentration on amplification performance. f) Comparison of the tolerances of different PCR systems for sDNA samples. GlaI‐P: GlaI‐digested 50 copies of methylated DNA per test; GlaI‐(P+sDNA): GlaI‐digested 50 copies of methylated DNA and 200 ng of sDNA; GlaI‐sDNA: GlaI‐digested 200 ng of sDNA. g) ColoC‐mSTEM PCR products separated by agarose gel electrophoresis line M:20 bp marker; line 1) 1000 copies per test; line 2) 100 copies per test; line 3) 40 copies per test; line 4) 10 copies per test; line 5) 10 ng undigested DNA per test. h) The sequence of ColoC‐mSTEM PCR products (78 bp of line 1 in F) detected by cloning sequencing.

Article Snippet: As expected, according to our multitarget analysis, the SDC2 gene or SDC2‐6 methylation site was always the most common factor due to the good clinical diagnostic performance of SDC2 gene methylation, especially at the SDC2‐6 methylation site.

Techniques: Amplification, Methylation, Modification, Concentration Assay, Comparison, Agarose Gel Electrophoresis, Marker, Sequencing, Cloning

Journal: Advanced Science

Article Title: Early Diagnosis of Colorectal Cancer Based on Bisulfite‐free Site‐specific Methylation Identification PCR Strategy: High‐Sensitivity, Accuracy, and Primary Medical Accessibility

doi: 10.1002/advs.202401137

Figure Lengend Snippet: The clinical performance of the 9 selected locations. a) Unsupervised hierarchical clustering of methylation markers differentially methylated between 67 normal and 80 CRC subjects in group 1. Each column represents an individual patient, and each row represents a methylation marker. b) Comparison of Ct values between normal controls and CRC patients in the training group. c) Receiver operating characteristic (ROC) curves and area under the curve values (AUCs) of the 5 loci in the SFRP2 gene. D) ROC curves and the AUCs of the 4 loci in the SDC2 gene. ** p < 0.01, *** p < 0.001.

Article Snippet: As expected, according to our multitarget analysis, the SDC2 gene or SDC2‐6 methylation site was always the most common factor due to the good clinical diagnostic performance of SDC2 gene methylation, especially at the SDC2‐6 methylation site.

Techniques: Methylation, Marker, Comparison

Journal: RSC Advances

Article Title: Morpholine-modified Ru-based agents with multiple antibacterial mechanisms as metalloantibiotic candidates against Staphylococcus aureus infection †

doi: 10.1039/d4ra02667e

Figure Lengend Snippet: (A) Antimicrobial morpholine contained agtents reported before; (B) the design of five Ru-based agents modified with morpholine and their synthesis routes.

Article Snippet: In this study, a series of ruthenium-based antibacterial agents modified with the morpholine moiety were designed and characterized, aiming to obtain a promising metalloantibiotic with a multitarget mechanism.

Techniques: Modification