Development of transformation for genome editing of an emerging model organism

Abstract

AbstractWith 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 flySciara (Bradysia) coprophila; this may serve as a paradigm for development of transformation for other emerging systems, especially insects.Sciarahas 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 ofSciararequires a transformation system to test mutations of DNA sequences that may play roles for these features. We describe aSciaratransformation system using a modified piggyBac transformation vector and detailed protocols we have developed to accommodateSciara-specific requirements. This advance will provide a platform for us and others in the growingSciaracommunity 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 Governor’s 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