Pqe 60 plasmid

The Escherichia coli JM109 competent cells, PrimeSTAR HS DNA polymerase, EmeraldAmp MAX PCR Master Mix, In-Fusion HD cloning kit, and NcoI were purchased from Takara Bio, Inc. (Shiga, Japan). The pQE-60 plasmid was obtained from Qiagen GmbH (Hilden, Germany). Poly(R)-3-hydroxybutyric acid, ( ±)3-hydroxybutyric acid (3HB) sodium salt and L-methionine sulfone were purchased from Sigma-Aldrich, Inc. (St. Louis, MO, USA). Acetone, sulfuric acid, methanol, sodium D-pantothenate, β-alanine, ampicillin sodium salt (Ap), chloramphenicol (Cm), isopropyl β-D-thiogalactopyranoside (IPTG), ethanol for HPLC, and chloroform for HPLC were purchased from FUJIFILM Wako Pure Chemical Corporation (Osaka, Japan). Oligonucleotides for PCR and DNA sequencing were purchased from Eurofins Genomics K.K. (Tokyo, Japan). PIPES sesquisogium and D-camphor-10-sulfonic acid were purchased from Dojindo Laboratories Co., Ltd. (Kumamoto, Japan) and Nacalai Tesque, Inc. (Kyoto, Japan), respectively. Amicon Ultra-0.5 Centrifugal Filter Units (3 kDa) were obtained from Merck KGaA (Darmstadt, Germany). All other chemicals were of reagent grade.

The PQE60 plasmid and E.Coli BL 21 were obtained from QIAGEN (Valencia, CA, USA). Isopropyl-1-thio-ß- D-galactopyranoside (IPTG) was from Bio-Basic Inc (Amherst, NY, USA) to induce recombinant SCF expression in bacteria. Guanidine Hydrochloride was from Amresco Inc (Solon, OH, USA). Urea was from Shanghai Experimental Reagent Inc (Shanghai, China). RPMI 1640 medium and STEM PRO®-34 SFM medium were from Gibco (Grand Island, NY, USA). Fetal bovine serum was from Hyclone (Logan, UT, USA). Goat-anti-mouse immunoglobulin G (IgG) conjugated with alkaline phosphatase was from Biolegend (Canada). Prestained protein molecular weight marker was from Bio-Rad (USA). Biotin conjugated goat anti-mouse-IgG was from Biolegend (San Diego, CA, USA). Mouse monoclonal antibody isotyping reagent kit and streptavidin-peroxidase was from Sigma-Aldrich (St. Louis, MO, USA). MACS immunomagnetic absorption column separation device and CD34 MicroBead Kit, was from Miltenyi Biotec (Germany). rhTPO and rhFLT-3 were from Pepro Tech (USA). Anti-human CD34 mAb conjugated with phycoerythrin was from Immunotec (Canada). Peptides were synthesized by Nanjing Genscript Inc. (Nanjing, China).