English (United Kingdom)

https://curated-unify.zendy.io/wp-json/zendy-region/v1/featured_content/oa?rat=en

https://curated-unify.zendy.io/wp-json/zendy-region/v1/highlighted_journal/

Global: Zendy Plus annual

Presents the access of premium content as premium feature

Premium Content

Presents the keyphrase highlighting as premium feature

Keyphrase Highlighting

Presents the summarisation as premium feature

Summarisation

Insights

Presents the pdf analysis as premium feature

PDF Analysis

Presents the zaia usage as premium feature

ZAIA

Global: Zendy Tools annual

Global: Zendy Free Plan

Global: Zendy Tools monthly

Global: Zendy Plus monthly

Genetic manipulation is an essential technique to analyze gene function; however, limited methods are available for  Babesia bovis , a causative pathogen of the globally important cattle disease, bovine babesiosis. To date, two stable transfection systems have been developed for  B .  bovis , using selectable markers blasticidin-S deaminase ( bsd ) or human dihydrofolate reductase ( hdhfr ). In this work, we combine these two selectable markers in a sequential transfection system. Specifically, a parent transgenic  B .  bovis  line which episomally expresses green fluorescent protein (GFP) and human dihydrofolate reductase (hDHFR), was transfected with a plasmid encoding a fusion protein consisting of red fluorescent protein (RFP) and blasticidin-S deaminase (BSD). Selection with WR99210 and blasticidin-S resulted in the emergence of parasites double positive for GFP and RFP. We then applied this method to complement gene function in a parasite line in which thioredoxin peroxidase-1 ( Bbtpx-1 ) gene was knocked out using hDHFR as a selectable marker. A plasmid was constructed harboring both RFP-BSD and  Bbtpx-1  expression cassettes, and transfected into a  Bbtpx-1  knockout (KO) parasite. Transfectants were independently obtained by two transfection methods, episomal transfection and genome integration. Complementation of  Bbtpx-1  resulted in full recovery of resistance to nitrosative stress, via the nitric oxide donor sodium nitroprusside, which was impaired in the  Bbtpx-1  KO parasites. In conclusion, we developed a sequential transfection method in  B .  bovis  and subsequently applied this technique in a gene complementation study. This method will enable broader genetic manipulation of  Babesia  toward enhancing our understanding of the biology of this parasite.

plos one

Transfection of Babesia bovis by Double Selection with WR99210 and Blasticidin-S and Its Application for Functional Analysis of Thioredoxin Peroxidase-1