This research amplified the phyA gene with the designed and synthesized primers specific for the phyA gene full-length coding sequence. The phyA gene was from Aspergillus niger F246 by the polymerase chain reaction(PCR), which is selected and identified in our laboratory. After sequncing the coding sequence, it was confirmed that the construction of cloning vector was succeeded. The phyA gene fragment was recovered from the pMD18T-phyA and ligated with prokaryotic expression vector pET30a+ to construct the recombinant expression plasmid pET30a+ -phyA. It was expressed with IPTG induction in E. coli for high efficiency. A new protein band with apparent molecular weight 50 kDa was detected in the lysate of the transformed cell by using SDS-PAGE. The amount of the soluble fusion protein was about 40% of large intestine bacillus soluble protein of transformed cells, estimated by absorbance scanning of SDS-PAGE and protein quantitation. It's phytase activity was eight times over the natural phyase. So this research provides the basis of the study on obtaining large and high active phytase and developmant of the new microbial ecologicalagent.
6-Phytase ; biosynthesis ; genetics ; Aspergillus niger ; enzymology ; genetics ; Base Sequence ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Genes, Fungal ; Genetic Vectors ; Molecular Sequence Data ; Recombinant Fusion Proteins ; biosynthesis ; genetics

OBJECTIVETo compare the short-term efficacy of laparoscope-assisted transanal total mesorectal excision (LA-taTME) and conventional laparoscopic TME (LTME) for rectal cancer by meta-analysis.

METHODSClinical studies that compared clinical outcomes of LA-taTME and LTME were searched from form PubMed, Embase, Ovid, CNKI and Wanfang database before January 2016. Two reviewers independently screened the articles and assessed the quality of the included studies by using the MINORS standard which involves 12 items. The score is 0-2 for each item and the maximum score is 24, and the ideal global score should be above16. RevMan 5.3 software was used for meta-analysis and outcome measures included operation time, hospital stay, number of harvested lymph node, rate of conversion, positive rate of circumferential resection margin and the rate of incomplete mesorectum.

RESULTSSeven studies were included in the analysis, and the score of all the studies was more than 16 points. A total of 479 patients (208 in LA-taTME, 271 in LTME) were enrolled. There were no significant differences in terms of age, sex, tumor location and clinical stage between two groups (all P>0.05). Results of meta-analysis showed that LA-taTME had lower rate of incomplete mesorectum (OR=0.29, 95% CI:0.10 to 0.84, P=0.02), lower rate of complications (OR=0.59, 95% CI:0.35 to 0.97, P=0.04) and shorter hospital stay (MD=-1.66, 95% CI:-3.22 to -0.11, P=0.04) than those of LTME, with significant differences. In terms of operation time (MD=-14.49, 95% CI:-37.87 to 8.90, P=0.22), number of harvested lymph node (MD=-0.45, 95% CI:-1.98 to 1.08, P=0.56), the rate of conversion (OR=0.31, 95% CI:0.08 to 1.24, P=0.10) and positive rate of circumferential resection margin (OR=0.43, 95% CI:0.17 to 1.04, P=0.06), there were no significant differences between two groups.

CONCLUSIONCompared to LTME, LA-taTME has similar short-term efficacy for rectal cancer, but it can reduce the rate of complications and rate of incomplete mesorectum.

Abdomen ; Digestive System Surgical Procedures ; methods ; Humans ; Laparoscopes ; Laparoscopy ; Length of Stay ; Operative Time ; Rectal Neoplasms ; surgery