CRISPR/Cas9-Mediated Homology-Directed Genome Editing in Pichia pastoris

2019 ◽  
pp. 211-225 ◽  
Author(s):  
Thomas Gassler ◽  
Lina Heistinger ◽  
Diethard Mattanovich ◽  
Brigitte Gasser ◽  
Roland Prielhofer
Keyword(s):  
Pichia Pastoris ◽  
Genome Editing

High efficiency CRISPR/Cas9 genome editing system with an eliminable episomal sgRNA plasmid in Pichia pastoris

2020 ◽  
Vol 138 ◽  
pp. 109556 ◽  
Author(s):  
Yankun Yang ◽  
Guoqiang Liu ◽  
Xiao Chen ◽  
Meng Liu ◽  
Chunjun Zhan ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Genome Editing ◽  
High Efficiency

A Novel and Efficient Genome Editing Tool Assisted by CRISPR-Cas12a/Cpf1 for Pichia pastoris

2021 ◽  
Author(s):  
Xinying Zhang ◽  
Songjie Gu ◽  
Xueyun Zheng ◽  
Siqi Peng ◽  
Yanru Li ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Genome Editing

Genome editing: Wollen wir das wirklich?

2017 ◽  
Vol 6 (3) ◽  
pp. 162-162
Author(s):  
Liane Kaufmann ◽  
Michael von Aster
Keyword(s):  
Genome Editing

Genome editing (CRISPR-Cas9) to identify and characterise functional variants determining metformin response

2018 ◽  
Author(s):  
M Keller ◽  
J Dalla-Riva ◽  
A Kurbasic ◽  
M Al-Majdoub ◽  
P Spegel ◽  
...  
Keyword(s):  
Genome Editing ◽  
Functional Variants

Optimization of fermentation conditions for the production of recombinant hpdgf-bb in Pichia pastoris

2015 ◽  
Vol 37 (1se) ◽  
Author(s):  
Duong Long Duy ◽  
Pham Minh Vu ◽  
Nguyen Tri Nhan ◽  
Tran Linh Thuoc ◽  
Dang Thi Phuong Thao
Keyword(s):  
Pichia Pastoris ◽  
Fermentation Conditions ◽  
Optimization Of Fermentation

Modification of the method for analysis of genome editing results using CRISPR/Cas9 system on preimplantation mouse embryos

2016 ◽  
pp. 15-20 ◽  
Author(s):  
T. V. Dimitireva ◽  
◽  
D. A. Reshetov ◽  
V. E. Zhernovkov ◽  
D. E. Vlodavets ◽  
...  
Keyword(s):  
Genome Editing ◽  
Mouse Embryos ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse

Cloning and Expression of Recombinant Human Insulin Gene in Pichia pastoris

2019 ◽  
Vol 10 (01) ◽  
Author(s):  
Rafid A. Abdulkareem
Keyword(s):  
Pichia Pastoris ◽  
Human Insulin ◽  
Expression Vector ◽  
Expression System ◽  
Insulin Gene ◽  
Cloning And Expression ◽  
Pcr Analysis ◽  
Major Band ◽  
Human Insulin Gene ◽  
Pichia Pastoris Expression System

The main goal of the current study was cloning and expression of the human insulin gene in Pichia pastoris expression system, using genetic engineering techniques and its treatment application. Total RNA was purified from fresh normal human pancreatic tissue. RNA of good quality was chosen to obtain a first single strand cDNA. Human preproinsulin gene was amplified from cDNA strand, by using two sets of specific primers contain EcoR1 and Notl restriction sites. The amplified preproinsulin gene fragment was double digested with EcoRI and Not 1 restriction enzymes, then inserted into pPIC9K expression vector. The new pPIC9K-hpi constructive expression vector was transformed by the heat-shock method into the E.coli DH5α competent cells. pPic9k –hpi, which was propagated in the positive transformant E. coli cells, was isolated from cells and then linearised by restriction enzyme SalI, then transformed into Pichia pastoris GS115 using electroporation method. Genomic DNA of His+ transformants cell was extracted and used as a template for PCR analysis. The results showed, that the pPic9k – hpi was successfully integrated into the P. pastoris genome, for selected His+ transformants clones on the anticipated band at 330 bp, which is corresponded to the theoretical molecular size of the human insulin gene. To follow the insulin expression in transformans, Tricine–SDS gel electrophoresis and Western blot analysis were conducted. The results showed a successful expression of recombinant protein was detected by the presence of a single major band with about (5.8 KDa) on the gel. These bands correspond well with the size of human insulin with the theoretical molecular weight (5.8 KDa).

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