Project description:The detection of hypermethylation markers on cell-free DNA (cfDNA) in biological fluids is a promising and non-invasive approach for early diagnosis and monitoring of human diseases. However, it is challenging to detect hypermethylation markers in a high-throughput, sensitive, and cost-effective manner. Here we presented a multiplex 5-methylcytosine marker barcode counting (MMBC-seq) technique and reported its clinical application for cfDNA from peripheral plasma samples. We identified an MMBC cancer detection panel and developed a scoring system to differentiate cancer versus healthy controls. In a multiple-cancer case-control study, the panel achieved a sensitivity and specificity of 80.2% and 95.7% respectively (AUC 0.906, 95% CI 0.846-0.948). The results suggest that MMBC-seq has great potential to realize non-invasive, flexible and clinically applicable cancer detection.
Project description:We developed a multiplex pseudo-isobaric dimethyl labeling (m-pIDL) method for proteome quantification to extend the capacity of the fragment ion-based method to 6-plex by one-step dimethyl labeling with several millidalton and dalton mass differences between precursor ions and enlarging the isolation window of precursor ions to 10 m/z during data acquisition.
Project description:The use of CRISPR/Cas proteins for the creation of multiplex genome-engineering represents an important avenue for crop improvement, and further improvements for creation of knock-in plant lines via CRISPR-based technologies may enable the high-throughput creation of designer alleles. To circumvent limitations of the commonly used CRISPR/Cas9 system for multiplex genome-engineering, we explored the use of Moraxella bovoculi 3 Cas12a (Mb3Cas12a) for multiplex genome-editing in Arabidopsis thaliana. We identified optimized promoter sequences for driving expression of single transcript multiplex crRNA arrays in A. thaliana, resulting in stable germline transmission of Mb3Cas12a-edited alleles at multiple target sites. By utilizing this system, we demonstrate single-transcript multiplexed genome-engineering using of up to 13 crRNA targets. We further show high target specificity of Mb3Cas12a-based genome-editing via whole-genome sequencing. Taken together, our method provides a simplified platform for efficient multiplex-genome-engineering in plant-based systems.