Synchronous online focus groups in health research: application and further development of methodology based on experiences from two mixed-methods research projects.

OBJECTIVES: Focus groups used for data collection in health research are increasingly conducted online. In two multi-center health research projects, we applied available methodological instructions for synchronous online focus groups (SOFGs). We describe necessary changes and specifications regarding the planning (recruitment, technology, ethics, appointments) and conduct (group composition, moderation, interaction, didactics) to enhance knowledge about the planning and conduct of SOFGs. RESULTS: Recruiting online proved to be challenging and necessitated direct and analogue recruiting, too. To ensure participation, less digital and more individual formats may be offered, e.g. telephone calls. Explaining verbally the specifics of data protection and anonymity in an online setting can foster participants' confidence to actively engage in the discussion. Two moderators, one moderating, one supporting technically, are advisable in SOFGs, however, due to limited nonverbal communication, roles and tasks need to be defined beforehand. Participant interaction is central to focus groups in general, but sometimes difficult to achieve online. Hence, smaller group size, sharing of personal information and moderators increased attention to individual reactions appeared helpful. Lastly, digital tools such as surveys and breakout rooms should be used with caution, as they easily inhibit interaction.

Isolation and characterization of new homing endonuclease specificities at individual target site positions.

Homing endonucleases are highly specific DNA endonucleases, encoded within mobile introns or inteins, that induce targeted recombination, double-strand repair and gene conversion of their cognate target sites. Due to their biological function and high level of target specificity, these enzymes are under intense investigation as tools for gene targeting. These studies require that naturally occurring enzymes be redesigned to recognize novel target sites. Here, we report studies in which the homodimeric LAGLIDADG homing endonuclease I-CreI is altered at individual side-chains corresponding to contact points to distinct base-pairs in its target site. The resulting enzyme constructs drive specific elimination of selected DNA targets in vivo and display shifted specificities of DNA binding and cleavage in vitro. Crystal structures of two of these constructs demonstrate that substitution of individual side-chain/DNA contact patterns can occur with almost no structural deformation or rearrangement of the surrounding complex, facilitating an isolated, modular redesign strategy for homing endonuclease activity and specificity.