Ni2 nta fast start kit

The expression vectors were transformed into Rosetta 2(DE3) competent cells (Novagen, Darmstadt, Germany) for CPP fusion protein expression. The recombinant proteins were purified using a Ni2⁺-NTA Fast Start Kit from QIAGEN (Hilden, Germany, Cat. # 30600). A centrifugal filter unit from Millipore (Bedford, MA, USA, Cat. # UFC501008) was applied to remove the salt and to concentrate purified proteins. Protein quantitation was performed with a Pierce™ BCA protein assay kit from Thermo Fisher (Waltham, MA, USA, Cat. # 23227). Endotoxin was removed with endotoxin removal beads from Miltenyi Biotec (Germany, Cat. # 130-093-659). Endotoxin levels were determined by using a Chromogenic LAL Endotoxin Assay Kit from GenScript (Nanjing, China, Cat. # L00350).

For the preparation of the nanocellulose carrier, sugar beet pulp (SBP) was provided by Sugar Beet Seed Institute (Karaj, Iran) and sulfuric acid (H₂SO₄, 98%) was purchased from Sigma–Aldrich. For enzyme assays, the 3, 5-dinitrosalicylic acid (DNS), bovine serum albumin (BSA), beechwood xylan, phenylmethane sulfonyl fluoride, imidazole, monosodium dihydrogen orthophosphate, metal ions, ethylene diaminetetra acetic acid (EDTA), urea, phenyl methyl sulfonyl fluoride (PMSF), sodium dodecyl sulfate (SDS), cetrimonium bromide (CTAB), tween 20, Triton X-100, Dithiothreitol (DTT), avicel, β-glucan, filter paper, locust bean gum (LBG), carboxymethyl cellulose (CMC) and β-pNPG were purchased from Sigma-Aldrich. The CRW was from the local market (Karaj, Iran). For the cloning, expression and purification of the PersiBGLXyn1, the T4 DNA ligase, NheI and SalI restriction enzyme and Gel Extraction kit were purchased from Thermo Fisher Scientific (Waltham, United States) and kanamycin (Duchefa, Haarlem, The Netherlands), Isopropyl β-D-1-thiogalactopyranoside (IPTG, Sigma–Aldrich), the Luria-Bertani medium (LB broth, Merck, Germany) and Ni2 + -NTA Fast Start Kit (Qiagen, Hilden, Germany) were used.