Ampflstr identifiler pcr amplification kit

Human soft tissue sarcoma cell lines were obtained from American Type Culture Collection (ATCC), Korean Cell Line Bank (KCLB), and other laboratories detailed in Additional file 1: Table S1. Each cell line was grown in appropriate culture medium (Additional file 1: Table S2) with 10% fetal bovine serum (Gibco, 16000-044) and 1% antibiotic–antimycotic 100× (Gibco, 15240-112). Cell lines were tested and validated for mycoplasma detection and human cell line authentication (STR DNA profiling) using AmpFLSTR™ Identifiler PCR Amplification Kit (Thermo Fisher Scientific, 4322288). For IFN-γ treatment, each cell line was seeded into six-well plates and treated with IFN-γ (R&D systems, 285-IF-100; 50 or 100 ng/ml) or BSA (Thermo Fisher Scientific, 23209; 50 or 100 ng/ml) as controls and incubated at 37 °C for 48 h.

Chidamide was supplied by Chipscreen Bioscience Ltd. (Shenzhen, China). ABT-199, Z-VAD-fmk, romidepsin, and vorinostat (SAHA) are all purchased from MedChemExpress (New Jersey, USA).
Molm-13 cells were purchased from AddexBio (San Diego, USA). MV4;11 and NB4 cells were purchased from ATCC (Teddington, UK). OCI-AML2, OCI-AML3 cells were kindly provided by Prof. Bing Z Carter (MD Anderson Cancer Center, USA). All cells were tested and authenticated by an AmpFlSTR Identifiler PCR Amplification Kit (Thermofisher Scientific, USA) in the year of 2018 in our laboratory, and were monthly tested for mycoplasma using PCR method. Peripheral blood samples of healthy donors for hematopoietic stem cell transplantation (n = 11) and bone marrow samples of patients with AML (n = 36) were obtained from the First Affiliated Hospital of Xiamen University with the informed consent for research purposes only. This study was performed in accordance with the Declaration of Helsinki and approved by the Ethics Review Board of First Affiliated Hospital of Xiamen University.

All cell lines were grown in media supplemented with 100 U ml⁻¹ penicillin, 100 μg ml⁻¹ streptomycin and 10% FBS and maintained in a 37 °C incubator with 5% CO₂. HK2 (ATCC CRL-2190), SN12-C (NCI-60), UO-31 (NCI-60), ACHN (ATCC CRL-1611) and MDA-MB-231 (ATCC HTB-26) were grown in DMEM. 786-O (ATCC CRL-1932) was grown in RPMI. For hypoxia, dishes were placed in an airtight chamber and flushed with 1% O₂, 5% CO₂ daily and placed in a 37 °C incubator50 51 (link). Cells were authenticated using the AmpFLSTR Identifiler PCR Amplification Kit (ThermoFisher). All cultures tested negative for mycoplasma contamination using the MycoAlert Mycoplasma Detection Kit (Lonza).

To generate regorafenib-resistant cell lines, HCT-116 and SW480 parental cells were exposed to regorafenib at the starting concentration of 3 µM and 5.5 µM, respectively, and continuously treated for four weeks. Then, an increasing regorafenib concentration (by steps of 0.5 µM) up to 6 µM and 7 µM for HCT-116 and SW480 cell lines was applied for six months. Cells were kept under the last medium conditions for another six months (Rego-12m). Cell Line Authentication was performed using STR (Short Tandem Repeat) profiling with AmpFLSTR™ Identifiler™ PCR Amplification Kit (Thermo Fisher Scientific, Waltham, MA, USA). DNA isolation was conducted from a cell pellet of 10⁶ cells and 16 independent PCR-systems were investigated and analysed (Eurofins Genomics, Ebersberg, Germany) (Supplementary Table S2).