ABSTRACT
BACKGROUND/AIM: The aim of this study was to evaluate N-acetylgalactosamine-6-sulfatase (GALNS) as a new biomarker candidate for detecting lung cancer. Glycodelin or PAEP, the serum levels of which are known to be elevated in lung and other cancers, served as a benchmark for comparison. PATIENTS AND METHODS: A total of 170 serum samples from healthy controls and patients with pneumonia, lung cancer, breast cancer, colon cancer, liver cancer, and head and neck cancer were analyzed for the levels of GALNS and PAEP by ELISA. RESULTS: The median serum levels of GALNS and PAEP in all cancer types as well as pneumonia patients were significantly higher than those of the healthy controls. CONCLUSION: In addition to previously known cancers, the median serum levels of PAEP were also found to be higher in liver and head and neck cancer patients. GALNS and PAEP are promising general biomarkers for multiple cancers and deserve further evaluation.
Subject(s)
Biomarkers, Tumor/blood , Chondroitinsulfatases/blood , Glycodelin/blood , Lung Neoplasms/blood , Area Under Curve , Benchmarking , Breast Neoplasms/blood , Case-Control Studies , Cell Line, Tumor , Colonic Neoplasms/blood , Enzyme-Linked Immunosorbent Assay , Female , Head and Neck Neoplasms/blood , Humans , Liver Neoplasms/blood , Lung/metabolism , Lung Neoplasms/diagnosis , Male , Pneumonia/bloodABSTRACT
Metabolic engineering often necessitates chromosomal integration of multiple genes but integration of large genes into Escherichia coli remains difficult. CRISPR/Cas9 is an RNA-guided system which enables site-specific induction of double strand break (DSB) and programmable genome editing. Here, we hypothesized that CRISPR/Cas9-triggered DSB could enhance homologous recombination and augment integration of large DNA into E. coli chromosome. We demonstrated that CRISPR/Cas9 system was able to trigger DSB in >98% of cells, leading to subsequent cell death, and identified that mutagenic SOS response played roles in the cell survival. By optimizing experimental conditions and combining the λ-Red proteins and linear dsDNA, CRISPR/Cas9-induced DSB enabled homologous recombination of the donor DNA and replacement of lacZ gene in the MG1655 strain at efficiencies up to 99%, and allowed high fidelity, scarless integration of 2.4, 3.9, 5.4, and 7.0 kb DNA at efficiencies approaching 91%, 92%, 71%, and 61%, respectively. The CRISPR/Cas9-assisted gene integration also functioned in different E. coli strains including BL21 (DE3) and W albeit at different efficiencies. Taken together, our methodology facilitated precise integration of dsDNA as large as 7 kb into E. coli with efficiencies exceeding 60%, thus significantly ameliorating the editing efficiency and overcoming the size limit of integration using the commonly adopted recombineering approach. Biotechnol. Bioeng. 2017;114: 172-183. © 2016 Wiley Periodicals, Inc.
