Transgene expression and silencing in a tick cell line: A model system for functional tick genomics.

Insect Biochemistry and Molecular Biology
T J KurttiU G Munderloh

Abstract

The genome project of the black legged tick, Ixodes scapularis, provides sequence data for testing gene function and regulation in this important pathogen vector. We tested Sleeping Beauty (SB), a Tc1/mariner group transposable element, and cationic lipid-based transfection reagents for delivery and genomic integration of transgenes into I. scapularis cell line ISE6. Plasmid DNA and dsRNA were effectively transfected into ISE6 cells and they were successfully transformed to express a red fluorescent protein (DsRed2) and a selectable marker, neomycin phosphotransferase (NEO). Frequency of transformation was estimated as 1 transformant per 5000-10,000 cells and cultures were incubated for 2-3 months in medium containing the neomycin analog G418 in order to isolate transformants. Genomic integration of the DsRed2 transgene was confirmed by inverse PCR and sequencing that demonstrated a TA nucleotide pair inserted between SB inverted/direct repeat sequences and tick genomic sequences, indicating that insertion of the DsRed2 gene into the tick cell genome occurred through the activity of SB transposase. RNAi using dsRNA transcribed from the DsRed2 gene silenced expression of red fluorescent protein in transformed ISE6 cells. SB tran...Continue Reading

References

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Citations

Jul 10, 2012·Experimental & Applied Acarology·Gerald BarryLesley Bell-Sakyi
Apr 14, 2010·Transgenic Research·Holly J FergusonMalcolm Fraser
Jul 21, 2015·Experimental & Applied Acarology·Erik Machado-FerreiraCarlos A G Soares
Apr 22, 2015·Experimental & Applied Acarology·Jonathan D OliverUlrike G Munderloh
Oct 5, 2016·Critical Reviews in Biochemistry and Molecular Biology·Suneel A NarayanavariZsuzsanna Izsvák
Dec 4, 2013·Applied and Environmental Microbiology·Jonathan D OliverUlrike G Munderloh
Nov 19, 2011·Applied Microbiology and Biotechnology·Vipin GopinathK Madhavan Nampoothiri
Jul 27, 2010·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Benjamin FränzelDirk Andreas Wolters
Aug 12, 2020·Scientific Reports·Kateryna KotsarenkoLibor Grubhoffer
Nov 14, 2020·Parasite Immunology·Sukanya NarasimhanErol Fikrig
May 28, 2021·Frontiers in Cellular and Infection Microbiology·Andrew NussMonika Gulia-Nuss

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