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Figure 5 | BMC Molecular Biology

Figure 5

From: Efficient four fragment cloning for the construction of vectors for targeted gene replacement in filamentous fungi

Figure 5

The constructed vectors for USER Friendly cloning. The constructed USER vector (A) pRF-HU2 (6323 bp), (B) pRF-HU2E (8696 bp), (C) pRF-HU (6336 bp) and (D) pRF-HUE (8709 bp). pRF-HU2 and pRF-HU2E were designed for single step directional cloning of two PCR amplicons to allow targeted gene replacement in filamentous fungi. The pRF-HU and pRF-HUE vectors were designed for construction of vectors for random integration into the fungal genome by non-homologous recombination. The LB UCS is identical in all four vectors, and the RB UCS is identical in the vectors for two fragment cloning. This allows for the reuse of primers with all four vectors. LB = left border, pTrpC = Tryptophan promoter form Aspergillus nidulans, hph = hygromycin phosphor transferase, TtrpC = Tryptophan terminator from A. nidulans, RB = right border, oriV = origin of replication in E. coli, KanR = kanamycin resistance, TrfA = replication initiation gene (broad-host-range), PgpdA = glyceraldehyde-3-phosphate dehydrogenase promoter from A. nidulans. Forward primers: RF-1 and RF-3; Reverse primers: RF-2.

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