Cloning, expression and purification of fructose-2, 6-bisphosphatase gene CpF2KP in papaya / 生物工程学报

Papaya, which is mainly cultivated in the southeastern region of China, is one of the four famous fruits in Lingnan. It is favored by people because of its edible and medicinal value. Fructose-6-phosphate, 2-kinase/fructose-2, 6-bisphosphatase (F2KP) is a unique bifunctional enzyme with a kinase domain and an esterase domain that catalyzes the synthesis and degradation of fructose-2, 6-bisphosphate (Fru-2, 6-P2), an important regulator of glucose metabolism in organisms. In order to study the function of the gene CpF2KP encoding the enzyme in papaya, it is particularly important to obtain the target protein. In this study, the coding sequence (CDS) of CpF2KP, with a full-length of 2 274 bp, was got from the papaya genome. The amplified sequence of full-length CDS was cloned into the vector PGEX-4T-1 which was double digested with EcoR I and BamH I. The amplified sequence was constructed into a prokaryotic expression vector by genetic recombination. After exploring the induction conditions, the results of SDS-PAGE showed that the size of the recombinant GST-CpF2KP protein was about 110 kDa. The optimum IPTG concentration and temperature for CpF2KP induction were 0.5 mmol/L and 28 ℃, respectively. The purified sin[A1] gle target protein was obtained after purifying the induced CpF2KP protein. In addition, the expression level of this gene was detected in different tissues, and showed that the gene was expressed at the highest level in seeds and the lowest in pulp. This study provides an important basis for further revealing the function of CpF2KP protein and studying the involved biological processes of this gene in papaya.

Clinical study on the efficacy and safety of percutaneous endoscopic anterior cervical discectomy in the treatment of cervical disc herniation.

PURPOSE: To compare the clinical efficacy of anterior percutaneous endoscopic cervical discectomy (APECD) and anterior cervical decompression and fusion (ACDF) in the treatment of cervical disc herniation. METHODS: A total of 115 cases of cervical disc herniation treated in our hospital from May 2016 to August 2018 were selected. The subjects were divided into APECD and ACDF groups. The operation time, intra-operative blood loss, and hospital stay of the two groups were recorded and compared, and the clinical symptoms were recorded at one week, one year, and two years after the operation. The neck and upper limb visual analog scale (VAS) score, Japanese Orthopedic Association (JOA) score, cervical range of motion (ROM), post-operative cervical lordosis angle and adjacent vertebral body height were compared between the two groups. RESULTS: All 115 patients underwent successful operations. The intra-operative blood loss and operation time in the APECD group were significantly less than those in the ACDF group (P < 0.05). There were no oesophago-tracheal sheaths, carotid artery sheaths, cervical sympathetic trunks, vertebral artery injuries, or dural tears in the two groups. The JOA and VAS scores of the two groups significantly improved from pre- to post-operatively, but there were no significant differences between the two groups at the last follow-up (P > 0.05). The post-operative radiological results showed that the herniated cervical intervertebral disc was removed completely. One year after the operation, the cervical lordosis angle increased significantly in both groups (P < 0.05). However, ROM decreased significantly in the ACDF group (P < 0.05), and there was no significant change in ROM in the APECD group (P > 0.05). The cervical lordosis angle did not significantly differ between the two groups at the last follow-up (P > 0.05), but there was a significant difference in ROM between the two groups at the last follow-up (P < 0.05). In the ACDF group, the height of the adjacent vertebral body increased 1 year after the operation (P < 0.05). In the APECD group, the height of the adjacent vertebral body decreased one year after the operation, but the value was not significantly different from the pre-operative value (P > 0.05). There was a significant difference in the height of the adjacent vertebral body between the two groups at the last follow-up (P < 0.05). The incidence of post-operative complications did not significantly differ between the two groups (P > 0.05). CONCLUSION: APECD is a minimally invasive treatment that is safe and effective. The medium- and short-term effects of APECD and ACDF in the treatment of cervical disc herniation are similar. In addition, APECD is superior to ACDF in the operation time, intra-operative blood loss, and ROM. However, prospective, multicenter studies with longer follow-up periods need to be conducted, and the recurrence rate and intervertebral height loss need to be studied.

Trans-interlamina percutaneous endoscopic cervical discectomy for symptomatic cervical spondylotic radiculopathy using the new Delta system.

To describe the rationale and surgical technique and compare the clinical effect of posterior percutaneous endoscopic cervical discectomy (PPECD) using the Delta system versus that of conventional PPECD (key-hole) surgery for the treatment of symptomatic cervical spondylotic radiculopathy (CSR). A retrospective analysis was performed on 106 single-segment CSR patients between February 2016 and February 2017, 50 of whom underwent conventional PPECD (key-hole), and 56 underwent PPECD using the Delta system. The operative time, intraoperative blood loss, intraoperative complications and postoperative hospital stay were recorded, and the clinical effect was evaluated by the indicators of the Neck Disability Index (NDI), arm-visual analog scale (arm-VAS), neck-VAS, EQ-5D and MacNab classification at the last follow-up. All patients underwent the operation successfully, and 106 patients were followed up. The operative time of the Delta group was 60.47 ± 0.71 min, while the operative time of the key-hole group was 75.46 ± 0.41 min. The difference between the two groups was statistically significant (P < 0.05). However, there was no significant difference between the two groups in terms of blood loss and hospital stay (P > 0.05). The VAS, NDI and EQ-5D scores of the neck and upper limbs in the two groups were significantly better than those before surgery at 1 week after surgery and at the last follow-up (P < 0.05). However, there was no significant difference between the two groups at the last follow-up (P > 0.05). At the last follow-up, there was no significant difference between the two surgical methods when evaluated using the modified MacNab criteria. The imaging results showed that the herniated disc was removed completely and the nerve root was decompressed. The complication rate in the Delta group (3/56, 5.35%) was significantly lower than that in the conventional key-hole group (5/50, 10.0%). PPECD using the Delta system for CSR may be a feasible and promising alternative surgical plan. Compared with the traditional key-hole method, this surgical system can not only provide the surgeon with a larger surgical field of vision but also reduces the operation time and complication rates.

"U" route transforaminal percutaneous endoscopic thoracic discectomy as a new treatment for thoracic spinal stenosis.

PURPOSES: To describe the rationale, surgical technique, and short-term follow-up results of a new minimally invasive treatment for thoracic spinal stenosis (TSS) caused by herniation, ossification of the ligamentum flavum (OLF), and/or ossification of the posterior longitudinal ligament (OPLL) with a "U" route transforaminal percutaneous endoscopic thoracic discectomy (PETD). METHODS: Fourteen patients, including seven males and seven females, underwent "U" route PETD. Myelopathy was caused by OLF in 14 patients, OPLL in one, combined OLF-OPLL in ten, and intervertebral disc herniation (IDH) in five. Decompression was performed in one segment in 12 patients, and in two segments in two patients. The Japanese Orthopedic Association (JOA) scores, visual analog scale (VAS) scores, and complications were documented. RESULTS: The JOA scores improved from 4.64 ± 2.31 pre-operatively to 7.07 ± 1.59 one day post-operatively and 11.79 ± 1.85 at final follow-up. The difference between pre-operation and post-operation was statistically significant (P < 0.05). Moreover, the VAS score was 6.07 ± 2.06 points pre-operatively, decreasing to 3.00 ± 1.24 points at one day post-operatively, and 1.14 ± 0.86 points at last follow-up (P < 0.05). Dural tear was observed in two cases during the intervention. No patient had transient worsening of pre-operative paralysis. CONCLUSIONS: This retrospective analysis shows that "U" route PETD for decompression may be a feasible alternative to treat thoracic spinal stenosis.