According to the amino acids sequence of OC-IdeltaD86 gene and Escherichia coli codon usage, we synthesized this gene by overlap extension PCR method with 7 oligonucleotides DNA fragments. The PCR fragment was inserted into pGEM-T-easy vector and the recombined plasmid was named pGEM-T-OC-IdeltaD86. Two oligonucleotides into which the BamH I and Xho I sites were introduced were designed and synthesized based on pGEM-T-OC-IdeltaD86 and pet21b, and the PCR fragment into which the BamH I and Xho I sites were introduced was obtained. After digesting it with BamH I and Xho I, OC-IdeltaD86 gene was cloned into the corresponding sites of pet21b and obtained prokaryotic expression vector pet21b-OC-IdeltaD86. OC-IdeltaD86 gene was expressed in E. coli (BL21(DE3)plysS) after IPTG(Isopropyl beta-D-1-thiogalactopyranoside) inducement for 5 hours. The fusion protein of OC-IdeltaD86:6His gene accounted for 11.4% of total protein and 16.4% of soluble protein, which had been successfully purified by Ni-NTA and concentrated by PEG20000. This protein can effectively inhibit papain activity in vitro and may be used in anti-nematode research in vivo.
Cloning, Molecular ; Cystatins ; biosynthesis ; genetics ; Cysteine Endopeptidases ; metabolism ; Cysteine Proteinase Inhibitors ; genetics ; Escherichia coli ; genetics ; metabolism ; Genes, Plant ; genetics ; Mutation ; Oligonucleotides ; chemical synthesis ; genetics ; Oryza ; genetics ; Papain ; antagonists & inhibitors ; Prokaryotic Cells ; metabolism ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; metabolism

Objective:To explore effects of implementing resistance exercise combined with Parkinson rehabilitation exercise in middle-aged and elderly patients with Parkinson disease accompanied by sleep disorder.Methods:By the convenient sampling method, 100 patients with Parkinson disease accompanied by sleep disorder who were hospitalized in Beijing Tiantan Hospital of Capital Medical University from August 2017 to August 2018 were selected as research objects. A total of 50 patients who were admitted to department of neurology were set as the control group and they were given conventional nursing. During the same period, a total of 50 patients who were admitted to department of neurology of the International Department were set as the observation group. On the basis of the control group, the observation group used resistance exercise combined with Parkinson rehabilitation exercises for intervention nursing. Pittsburgh Sleep Quality Index (PSQI) and Parkinson's Disease Quality of Life Questionnaire (PDQ-39) were used to evaluate the sleep quality and quality of life of patients in the two groups before intervention and 4 weeks after the intervention.Results:After intervention, the total PSQI score of the observation group was (20.76±7.71) , the subjective sleep quality dimension score was (1.24±0.80) , the time for falling asleep dimension score was (2.55±1.50) , the sleep time dimension score was (1.31±1.08) , the sleep efficiency dimension score was (1.56±1.29) , the sleep disorder dimension score was (6.07±4.38) , the hypnotic drug dimension score was (1.20±1.18) and the daytime dysfunction dimension score was (2.78±1.83) , which were lower than those of the control group. There were statistically significant differences in the total score of PSQI, sleep disorder dimension and daytime dysfunction dimension ( P<0.05) . The quality of life score of the observation group after intervention was (120.86±21.98) , lower than that of the control group before the intervention, the differences were statistically significant ( P<0.05) . Conclusions:Resistance exercise combined with Parkinson rehabilitation exercise has a certain effect on improving sleep quality and quality of life of patients with Parkinson accompanied by sleep disorder. The operation method is simple and safe, and it is worthy of clinical promotion and application.