Download Glip Apk PORTABLE
Keiko Bludworth
To construct the Δaspf1 ΔgliP double mutant, 1352 bp of the aspf1 5ʹ flanking sequence was PCR-amplified from Af293 genomic DNA using primers Aspf1-BLE-F4 and Aspf1-BLE-F3 (Table S3) and 1413 bp of the aspf1 3ʹ-flanking sequence was PCR-amplified using primers Aspf1-BLE-F4 and Aspf1-BLE-F3. A DNA fragment containing the phleomycin resistance gene (ble) was PCR-amplified from plasmid p402 [48] using primers BLE-F and BLE-R (Table S3). Next, a DNA fragment containing the aspf1 5ʹ flanking region linked to the 5ʹ portion of ble obtained by fusion PCR using primers Aspf1-BLE-F4 and BL (Table S3). A fragment containing the aspf1 3ʹ flanking region linked to the 3ʹ portion of ble was obtained similarly using primers Aspf1-BLE-F1 and LE (Table S3). These two fragments were used to transform the A. fumigatus Δglip mutant [16]. Phleomycin resistant clones were screened for deletion of aspf1 by colony PCR using primers Aspf1-RT-F and Aspf1-RT-R.