Generation of transgenic zebra finches by the culture and genetic modification of germline stem cells

Abstract

Genes are DNA segments that encode the instructions to make particular proteins or RNA molecules (Alberts et al., 2008). They are the units of heredity and together with the environment they shape the functioning and phenotype of all living creatures. Small alterations in the expression of a gene can have implications in the performance of complex networks connecting DNA, RNA and proteins leading to adaptation or disfunction. Therefore, experimental methods to modify gene expression are essential to reach a deeper knowledge of the gene’s function. Animal species with structural, functional or developmental similarities to man are used with great success to study gene function, for example in the field of human neurodegenerative diseases. Different methods were developed to produce animal models with an alteration in the expression of a particular gene to comprehend the mechanisms behind genetic diseases and to come up with gene therapies to cure them. Songbirds in particular are commonly used as an animal model to study the genetic and neural basis of vocal learning and communication, as well as sex hormone dependent brain development and adult neurogenesis (Goldman and Nottebohm, 1983; Paton and Nottebohm, 1984; Gahr, 2007; Balthazart et al., 2010; McCarthy and Arnold, 2011; Mello, 2014; Prather et al., 2017; Mooney, 2020). Songbirds are vocal learners that possess many parallels with speech acquisition in humans and therefore constitute an excellent animal model to study speech and language disorders (Abe et al., 2015; Liu et al., 2015). In the scope of this thesis I have developed a new method to produce transgenic songbirds by the culture and genetic modification of germline stem cells (Gessara et al., 2021), since genetic manipulation in songbirds has not yet worked routinely. For this it was necessary to examine and optimize the genetic signalling pathways and biological factors of the culture of primordial germ cells. To prepare a first application of my transgenic technology, I focused on the song control system, responsible for learning and production of song in songbirds. I have studied the structure and function of physiological variations of the androgen receptor and propose a method for the genetic alteration of the androgen receptor in a zebra finch transgenic model to further investigate its role in song learning and development.