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bioRxiv · 10.1101/2022.04.19.488772

Development of transformation for genome editing of an emerging model organism

Yamamoto, Y.·Gerbi, S. A.
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Abstract

With the advances in genomic sequencing, many organisms with novel biological properties are ripe for use as emerging model organisms. However, to make full use of them, transformation methods need to be developed to permit genome editing. Here, we present development of transformation for the fungus fly Sciara (Bradysia) coprophila; this may serve as a paradigm for development of transformation for other emerging systems, especially insects. Sciara has a variety of unique biological features including locus-specific developmentally-regulated DNA amplification; chromosome imprinting; a monopolar spindle in male meiosis I; non-disjunction of the X chromosome in male meiosis II; X chromosome elimination in early embryogenesis; germ line limited (L) chromosomes; high resistance to radiation. Mining the unique biology of Sciara requires a transformation system to test mutations of DNA sequences that may play roles for these features. We describe a Sciara transformation system using a modified piggyBac transformation vector and detailed protocols we have developed to accommodate Sciara-specific requirements. This advance will provide a platform for us and others in the growing Sciara community to take advantage of this unique biological system. In addition, the versatile piggyBac vectors described here and transformation methods will be useful for other emerging model systems. Author BiographiesSusan A. Gerbi (Ph.D. with Joseph Gall at Yale University 1970) is the George Eggleston, Professor of Biochemistry at Brown University. Her research includes chromosomes, DNA replication and ribosomal RNA. She was President and is a Fellow of ASCB, a Fellow of AAAS and received the RI Governors Award for Scientific Achievement. Other honors include RNA Society/CSHL Press Distinguished Research Mentor award; GSA George Beadle award; ASCB Senior Leadership/ Mentoring Award. She is a national leader in graduate education, including member of the National Academy of Sciences Panel on Bridges to Independence that led to the NIH K99 program, Chair of the AAMC Graduate Research Education Training Group; Chair of the FASEB Consensus Conference on Graduate Education. Yutaka Yamamoto (M.D. Kansai Medical University 1990; Ph.D. with Walter Gehring at Biozentrum - Basel 1995; postdoc with David Glover at Dundee University and University of Cambridge) is a research associate at Brown University. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=65 SRC="FIGDIR/small/488772v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@17fce9aorg.highwire.dtl.DTLVardef@1d4e96forg.highwire.dtl.DTLVardef@129a689org.highwire.dtl.DTLVardef@1748fd8_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Yamamoto, Y., Gerbi, S. A.. 2022-04-19. Development of transformation for genome editing of an emerging model organism. https://doi.org/10.1101/2022.04.19.488772

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