Mapping of Integrations

Shawn M. Burgess, National Human Genome Research Institute, NIH | external teacher
Melanie König, Hiroshi Kikuta | local teaching assistants

A major question in the study of retroviruses and transposable elements as genetic tools is whether the integrations occur randomly throughout the genome or have specific sequence or structural preferences. This question is relevant to understanding the mechanism of integration for the various elements, but it is also of critical importance when using these various insertional elements as a mutagen or as a tag for genetic analysis. By understanding what biases the elements may possess, it will allow the researcher to determine how many genes can actually be disrupted by a particular element, and how many integrations it would take to reach "saturation" mutagenesis.
The completion of the human genome project (and the advanced state of other genomes) in combination with newly developed techniques have made it possible to determine how random these various insertional elements are. The high throughput isolation of flanking sequence we developed for the human genome can in principle also applied to the zebrafish genome to analyze insertions in this course. We will cover what has been learned about several insertional elements including Murine Leukemia Virus based vectors (MLV), HIV-1 based vectors, the sleeping beauty transposon, and other commonly used elements. We will also discuss computational methods used to determine what kinds of integration preferences these elements might have.

Suggested Readings

  • Transgenic zebrafish produced by retroviral infection of in vitro cultured sperm, PNAS In Press.

  • Nissen R, Yan J, Amsterdam A, Hopkins N, and Burgess S. (2003) Zebrafish foxi one modulates cellular responses to fgf signaling required for the integrity of ear and jaw patterning, Dev. 130, 2543-2554.

  • Wu X, Li Y, Crise B, Burgess S. (2003) Transcription start regions in the human genome are favored targets for MLV integration, Science 300, 1749-1751.

  • Chen, W, Burgess, S, Golling, G, Amsterdam, A, and Hopkins, N. (2002) High-Throughput Selection of Retrovirus Producer Cell Lines Leads to Markedly Improved Efficiency of Germ Line-Transmissible Insertions in Zebrafish, J. Vir. 76, 2192-2198.

  • Golling G, Amsterdam A, Sun Z, Antonelli M, Maldonado E, Chen W, Burgess S, Haldi M, Artzt K, Farrington S, Lin SY, Nissen RM, and Hopkins N (2002) Insertional mutagenesis in zebrafish rapidly identifies genes essential for early vertebrate development, Nat. Genet. 31, 135-140.