A general method to isolate genes tagged by a high copy number transposable element

Area of Science:

• Plant genetics and molecular biology
• Transposable element research
• Gene tagging and isolation

Background:

• The Petunia hybrida line W138 harbors over 200 copies of the transposable element dTph1.
• This high copy number leads to frequent, unstable mutations in W138 progeny.
• Efficient methods are needed to isolate genes affected by these transposable elements.

Purpose of the Study:

• To develop a method for isolating dTph1 flanking sequences specific to mutant plants.
• To create a probe for identifying mutated genes in Petunia.
• To assess the general applicability of the method for other transposable elements.

Main Methods:

• Utilized inverse polymerase chain reaction (IPCR) to amplify dTph1 flanking sequences.
• Employed differential screening of cloned IPCR products to identify mutant-specific sequences.
• Generated probes from unique flanking sequences for library screening.

Main Results:

• Successfully isolated dTph1 flanking sequences unique to mutant Petunia plants.
• Developed a direct method to generate probes for mutated genes.
• Demonstrated the probe's utility in screening existing cDNA and genomic libraries.

Conclusions:

• The developed method efficiently isolates genes tagged by the high copy number transposable element dTph1.
• This approach provides a valuable tool for studying gene function and mutations in Petunia.
• The technique is potentially applicable to isolating genes tagged by other high copy number transposable elements in various plant species.