The formation of definitive endoderm (DE) is a fundamental step for the development of the gastrointestinal tract, respiratory tract and endocrine organs. We present a CRISPR-based pooled screening approach to identify genes which contribute to DE induction from hiPSCs in vitro. CRISPR-based pooled genetic screens in mammalian cell culture enable researchers to identify genes required for a cellular phenotype of interest in an unbiased way. To enable a CRISPR-based forward genetic screen for identifying regulatory genes required for DE differentiation from hiPSCs, we performed pooled screens using a human genome-scale CRISPR knockout library. In addition, we performed a transcriptional activation screen using a lentiviral CRISPRa library to identify the downstream targets of the TGF/nodal/activin signaling pathway, which is a key signaling pathway for DE specification. We identified several signaling pathways including TGFβ, Erk, JNK, and CREB pathways are involved with DE differentiation. We suggest that CRISPR-based pooled genetic screens are a useful tool to identify key signaling pathways and genes required for in vitro differentiation processes and are served as a platform to improve differentiation protocols.

Plant biomass is a huge carbon-complex that has potential as a nutrient. Therefore we extracted and separated useful materials for plant growth from tea leaf and stem. The pre-treatment process including high temperature (200 °C) and pressure (20-40 kgf/cm2) was treated for several minutes and extracted at 120 °C for 30-60 minutes. After that the chemical compositions and ingredients were analyzed from that plantnutrient. As a result of mineral contents, calcium and magnesium concentrations are higher than other minerals. Also the result of carbohydrates analyses has shown that the sugar oligomer consists of xylose(95.3%) and glucose(4.7%), and the sugar monomer consists in the order of xylose (52.7%) > manose (22.8%) > arabinose (10.8%) > galactose (10.2%) > glucose (3.5%). Before applied to field, in vitro plant growth system and formulation were examined. To evaluate the effect of the nutrients, both strawberry green-house and persimmon fields were used in this test. The treated persimmons were heavier than controls scored at 13-22%. In addition, the storageperiod was extended in the treated strawberries. Interestingly in the treated strawberry, the contents of polyphenols were increased (38-57%). These results suggest that the plant-nutrient can afford to help for plant growth and storage, and it can be substituted for other commercial nutrients. In conclusion, this plant-nutrient may help to extend eco-friendly or organic farming in Hadong-gun area.