PMID- 32493474
OWN - NLM
STAT- MEDLINE
DCOM- 20210304
LR  - 20231127
IS  - 1756-0500 (Electronic)
IS  - 1756-0500 (Linking)
VI  - 13
IP  - 1
DP  - 2020 Jun 3
TI  - A transient CRISPR/Cas9 expression system for genome editing in Trypanosoma 
      brucei.
PG  - 268
LID - 10.1186/s13104-020-05089-z [doi]
LID - 268
AB  - OBJECTIVE: Generation of knockouts and in situ tagging of genes in Trypanosoma 
      brucei has been greatly facilitated by using CRISPR/Cas9 as a genome editing 
      tool. To date, this has entailed using a limited number of cell lines that are 
      stably transformed to express Cas9 and T7 RNA polymerase (T7RNAP). It would be 
      desirable, however, to be able to use CRISPR/Cas9 for any trypanosome cell line. 
      RESULTS: We describe a sequential transfection expression system that enables 
      transient expression of the two proteins, followed by delivery of PCR products 
      for gRNAs and repair templates. This procedure can be used for genome editing 
      without the need for stable integration of the Cas9 and T7RNAP genes.
FAU - Shaw, Sebastian
AU  - Shaw S
AD  - Institute of Cell Biology, University of Bern, Bern, Switzerland.
AD  - Graduate School of Cellular and Biomedical Science, University of Bern, Bern, 
      Switzerland.
AD  - Department of Pathobiology, School of Veterinary Medicine, University of 
      Pennsylvania, 380 South University Avenue, Philadelphia, PA, 19104, USA.
FAU - Knusel, Sebastian
AU  - Knusel S
AD  - Institute of Cell Biology, University of Bern, Bern, Switzerland.
FAU - Hoenner, Sarah
AU  - Hoenner S
AD  - Institute of Cell Biology, University of Bern, Bern, Switzerland.
FAU - Roditi, Isabel
AU  - Roditi I
AUID- ORCID: 0000-0003-2812-6513
AD  - Institute of Cell Biology, University of Bern, Bern, Switzerland. 
      isabel.roditi@izb.unibe.ch.
LA  - eng
GR  - 310030_184669/Schweizerischer Nationalfonds zur Forderung der Wissenschaftlichen 
      Forschung/
GR  - 31003A_166427/Schweizerischer Nationalfonds zur Forderung der Wissenschaftlichen 
      Forschung/
PT  - Journal Article
DEP - 20200603
PL  - England
TA  - BMC Res Notes
JT  - BMC research notes
JID - 101462768
RN  - 0 (Viral Proteins)
RN  - EC 2.7.7.- (bacteriophage T7 RNA polymerase)
RN  - EC 2.7.7.6 (DNA-Directed RNA Polymerases)
RN  - 0 (RNA, Guide, Kinetoplastida)
SB  - IM
MH  - *CRISPR-Cas Systems
MH  - DNA-Directed RNA Polymerases/genetics
MH  - *Gene Editing
MH  - Genome, Protozoan/*genetics
MH  - RNA, Guide, Kinetoplastida/genetics
MH  - Trypanosoma brucei brucei/*genetics
MH  - Viral Proteins/genetics
PMC - PMC7268226
OTO - NOTNLM
OT  - CRISPR/Cas9
OT  - Transient transfection
OT  - Trypanosome
COIS- We, the authors, declare no conflict of interests.
EDAT- 2020/06/05 06:00
MHDA- 2021/03/05 06:00
PMCR- 2020/06/03
CRDT- 2020/06/05 06:00
PHST- 2020/04/04 00:00 [received]
PHST- 2020/05/13 00:00 [accepted]
PHST- 2020/06/05 06:00 [entrez]
PHST- 2020/06/05 06:00 [pubmed]
PHST- 2021/03/05 06:00 [medline]
PHST- 2020/06/03 00:00 [pmc-release]
AID - 10.1186/s13104-020-05089-z [pii]
AID - 5089 [pii]
AID - 10.1186/s13104-020-05089-z [doi]
PST - epublish
SO  - BMC Res Notes. 2020 Jun 3;13(1):268. doi: 10.1186/s13104-020-05089-z.