Abstract

P-element vectors are commonly used to make transgenic Drosophila and generally insert in the genome in a nonselective manner. However, when specific fragments of regulatory DNA from a few Drosophila genes are incorporated into P-transposons, they cause the vectors to be inserted near the gene from which the DNA fragment was derived. This is called P-element homing. We mapped the minimal DNA fragment that could mediate homing to the engrailed/invected region of the genome. A 1.6 kb fragment of engrailed regulatory DNA that contains two Polycomb-group response elements (PREs) was sufficient for homing. We made flies that contain a 1.5kb deletion of engrailed DNA (enΔ1.5) in situ, including the PREs and the majority of the fragment that mediates homing. Remarkably, homing still occurs onto the enΔ1. 5 chromosome. In addition to homing to en, P[en] inserts near Polycomb group target genes at an increased frequency compared to P[EPgy2], a vector used to generate 18,214 insertions for the Drosophila gene disruption project. We suggest that homing is mediated by interactions between multiple proteins bound to the homing fragment and proteins bound to multiple areas of the engrailed/invected chromatin domain. Chromatin structure may also play a role in homing.

Highlights

  • P-element mediated transformation has been used to generate transgenic Drosophila for about 30 years [1]

  • A 1.6kb fragment of en DNA can mediate homing Our previous results indicated that a 2.6 kb fragment of en DNA, extending from 22.4 kb upstream through +188bp of the en transcription unit could mediate P-element homing to the en/inv

  • We show that a 1.6kb fragment that extends from 22.0 kb through 20.4 kb is sufficient for homing

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Summary

Introduction

P-element mediated transformation has been used to generate transgenic Drosophila for about 30 years [1]. The first case of this was described by Hama, Ali, and Kornberg in 1990 [7]; regulatory DNA from the engrailed (en) gene, when included in a P-element vector (P[en]), caused P[en] to insert in the vicinity of the en gene at a high frequency. For homing mediated by en, bithorax, and eve DNA, insertions are not site-specific but are regional; that is, they occur in the vicinity of the endogenous gene but can be distributed over a large genomic region. For homing by P[en], insertions occur over about a 300 kb region, including en, inv and flanking genes [7,11,12]

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