No evidence of genome editing activity from Natronobacterium gregoryi Argonaute (NgAgo) in human cells

The argonaute protein from the thermophilic bacterium Thermus thermophilus shows DNA-guided DNA interfering activity at high temperatures, complicating its application in mammalian cells. A recent work reported that the argonaute protein from Natronobacterium gregoryi (NgAgo) had DNA-guided genome e...

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Bibliographic Details
Published in:PloS one 2017-05, Vol.12 (5), p.e0177444
Main Authors: Javidi-Parsijani, Parisa, Niu, Guoguang, Davis, Meghan, Lu, Pin, Atala, Anthony, Lu, Baisong
Format: Article
Language:English
Subjects:
Antigens
Argonaute Proteins - genetics
Argonaute Proteins - metabolism
Biology and Life Sciences
Blotting, Western
Data processing
Deoxyribonucleic acid
DNA
DNA sequencing
Editing
Gene Editing
Gene expression
Gene sequencing
Genetic aspects
Genome editing
Genome, Human - genetics
Genomes
Health sciences
HEK293 Cells
High temperature
High temperatures
High-Throughput Nucleotide Sequencing
Humans
Laboratories
Mammalian cells
Mammals
Medicine
Natronobacterium - genetics
Physiological aspects
Plasmid DNA
Plasmids
Plasmids - genetics
Proteins
Research and Analysis Methods
Staphylococcus aureus - genetics
Staphylococcus aureus - metabolism
Stem cells
Thermus thermophilus
Thermus thermophilus - genetics
Thermus thermophilus - metabolism
Transfection
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Summary:The argonaute protein from the thermophilic bacterium Thermus thermophilus shows DNA-guided DNA interfering activity at high temperatures, complicating its application in mammalian cells. A recent work reported that the argonaute protein from Natronobacterium gregoryi (NgAgo) had DNA-guided genome editing activity in mammalian cells. We compared the genome editing activities of NgAgo and Staphylococcus aureus Cas9 (SaCas9) in human HEK293T cells side by side. EGFP reporter assays and DNA sequencing consistently revealed high genome editing activity from SaCas9. However, these assays did not demonstrate genome editing activity by NgAgo. We confirmed that the conditions allowed simultaneous transfection of the NgAgo expressing plasmid DNA and DNA guides, as well as heterologous expression of NgAgo in the HEK293T cells. Our data show that NgAgo is not a robust genome editing tool, although it may have such activity under other conditions.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0177444