Mycoplasma hoechst stain kit

Maintenance of undifferentiated H9-hESCs and differentiation into M-MSCs (Fig. 1a) were performed as previously described^{23 (link), 24 (link)}. The established M-MSCs were cultured with EGM2-MV medium (Lonza, San Diego, CA, USA) on plates coated with rat tail collagen type I (Sigma-Aldrich, St. Louis, MO, USA) in a humidified atmosphere with 5% CO₂ at 37 °C. All M-MSCs used in experiments were expanded less than ten passages to ensure multipotency. Characterization of basic features such as surface protein expression, cell proliferation, multipotency (in vitro differentiation into osteogenic, chondrogenic, or adipogenic lineages), in vitro angiogenesis assays, and karyotyping were performed as previously described^{23 (link), 24 (link)}. The M-MSC line stably expressing GFP was established by infection of GFP-expressing lentivirus produced as previously described^{14 (link)}. Human BM-MSCs purchased from Lonza (Basel, Switzerland) were cultured following the manufacturer’s instructions. Cells expanded for fewer than seven passages were used for experiments to ensure multipotency. All cells were tested for mycoplasm content each month (Mycoplasma Hoechst Stain Kit; 3030000; MP Biomedicals, LLC, Santa Ana, CA, USA).

The strongly adherent P. xylostella Pxem_ZJU cell line, which was derived from the P. xylostella, was maintained in 60 mm culture dish in TNM-FH culture medium plus 10% fetal bovine serum (FBS) at 27 °C under ambient atmosphere^{38 (link)}. The human embryonic kidney cell 293 (HEK293) cell line was bought from Thermo Fisher and maintained at 37 °C in 5% CO₂ in DMEM basic medium (Gibco) medium plus 10% FBS. All cell lines were verified to be mycoplasma-free using the PCR-based mycoplasma detection kit (Kaiji, Nanjing, China). Mycoplasma testing was performed on all lines using Mycoplasma Hoechst Stain Kit (MP Biomedicals, Solon, OH, USA).