The recombinant allergens of P. aztecus, (Pen a 1) were purchased from Indoor Biotechnologies (Product Code: NA-STM-1) (Indoor Biotechnologies, Bangalore, India).
The cDNA of Der p 10 was synthesized using primer dT17 and MMLV reverse transcriptase (Invitrogen, Massachusetts, USA) from total mRNA of D. pteronyssinus. The specific primers of Der p 10 were designed based on the accession number, AF016278.150 (link), provided in the National Center for Biotechnology Information. The polymerase chain reaction (PCR) products of Der p 10 with six histidine (His) were ligated into a pQE30 vector and transformed into Escherichia. coli- M15. The selection was performed using kanamycin (25 μg/ml) and ampicillin (100 μg/ml). The 6 × His tagged recombinant proteins were induced using 1 mM isopropyl-β-D- thiogalactopyranoside (Promega, Wisconsin, USA). The proteins were purified by nickel-nitrilotriacetic acid agarose metal affinity column chromatography under native conditions. The results of purified rDer p 10 are shown in the Supplementary Fig.1 .
The cDNA of Der p 10 was synthesized using primer dT17 and MMLV reverse transcriptase (Invitrogen, Massachusetts, USA) from total mRNA of D. pteronyssinus. The specific primers of Der p 10 were designed based on the accession number, AF016278.150 (link), provided in the National Center for Biotechnology Information. The polymerase chain reaction (PCR) products of Der p 10 with six histidine (His) were ligated into a pQE30 vector and transformed into Escherichia. coli- M15. The selection was performed using kanamycin (25 μg/ml) and ampicillin (100 μg/ml). The 6 × His tagged recombinant proteins were induced using 1 mM isopropyl-β-D- thiogalactopyranoside (Promega, Wisconsin, USA). The proteins were purified by nickel-nitrilotriacetic acid agarose metal affinity column chromatography under native conditions. The results of purified rDer p 10 are shown in the Supplementary Fig.
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