ABSTRACT
Gluconobacter oxydans are widely used in industrial due to its ability of oxidizing carbohydrate rapidly. However, the limited gene manipulation methods and less of efficient gene editing tools impose restrictions on its application in industrial production. In recent years, the clustered regularly interspaced short palindromic repeats (CRISPR) system has been widely used in genome editing and transcriptional regulation which improves the efficiency of genome editing greatly. Here we constructed a CRISPR/dCpf1-mediated gene transcriptional repression system, the expression of a nuclease inactivation Cpf1 protein (dCpf1) in Gluconobacter oxydans together with a 19 nt direct repeats showed effective repression in gene transcription. This system in single gene repression had strong effect and the relative repression level had been increased to 97.9%. While it could be applied in multiplex gene repression which showed strong repression ability at the same time. Furthermore, this system was used in the metabolic pathway of L-sorbose and the regulatory of respiratory chain. The development of CRISPR transcriptional repression system effectively covered the shortage of current gene regulation methods in G. oxydans and provided an efficient gene manipulation tool for metabolic engineering modification in G. oxydans.
Subject(s)
CRISPR-Cas Systems/genetics , Clustered Regularly Interspaced Short Palindromic Repeats/genetics , Gene Editing , Gene Expression , Gluconobacter oxydans/genetics , Metabolic EngineeringABSTRACT
BACKGROUND: Glaucoma is a kind of eye disease that can cause irreversible blindness which is characterized by visual field loss. Clinical research shows that the optic nerve head has changed before the visual field loss. The morphological changes of the optic nerve head have become the key to determine the early diagnosis of glaucoma and disease development. So it has important significance for us to study the morphological changes of the tissues of optic nerve head under the high intraocular pressure. OBJECTIVE: To establish three-dimensional finite element model of optic nerve head which includes choroid, retina and lamina cribrosa, and analyze the thickness of the optic nerve head under the acute high intraocular pressure. METHODS: (1) We chose healthy cats without refractive media and other eye diseases. The tomographic sequence images of the optic nerve head of a cat were obtained with the intraocular pressure of the normal value by using enhanced depth imaging spectral-domain optical coherence tomography. (2) The three-dimensional model of the retina, choroid and lamina cribrosa was obtained. Three-dimensional model of optic nerve head was established by assembling the model of the retina, the choroid and the lamina cribrosa. The thickness of the retina, choroid and lamina cribrosa was analyzed under different intraocular pressures by using the method of finite element analysis. (3) Animal model of acute ocular hypertension was established by methods of anterior chamber perfusion. The tomographic sequence images of the optic nerve head of a cat were obtained with different intraocular pressures by using enhanced depth imaging spectral-domain optical coherence tomography. Then, we measured the thickness changes of the choroid, retina and lamina cribrosa under different intraocular pressures, and compared with the results of finite element analysis. RESULTS AND CONCLUSION: The thickness of the choroid, retina and lamina cribrosa trended to be thin, and the cup dish ratio of optic nerve head increased gradual y with the increased intraocular pressures. Regarding to the thickness changes of the choroid, retina and lamina cribrosa, the trend of experimental measurement results was consistent with finite element calculation results. Hence, it is feasible to analyze morphological changes of every fundus tissue under high intraocular pressure using the tomographic sequence images obtained by optical coherence tomography. We can predict the morphological changes of the optic nerve head tissue by finite element analysis, which has certain guiding significance in determining the progression of glaucoma.
ABSTRACT
Objective To evaluate the effect of phacoemulsification and intraocular lens implantation with topical anesthesia in out-patients department.Methods Phacoemulsification and intraocular lens implantation were performed successively in 132 patients(157 eyes) with topical anesthesia.Results Phacoemulsification and intraocular lens implantation were successfully performed in 152 eyes(96.8%)with topical anesthesia and 5 eyes(3.2%)need retrobulbar anesthesia additionally.The visual acuity and complications after operation in the topical anesthesia were the same as that performed previously under retrobular anesthesia.Conclusions Phacoemulsification and intraocular lens implantation may be performed with topical anesthesia through scleral tunnel incision.It is convenient for us to perform the operations on the out-patient patients with cataract.