![]() ![]() We show that our vector set is applicable for the biotechnologically relevant Penicillium chrysogenum and the developmental model system Sordaria macrospora. To make cloning most feasible, we provide robust protocols, namely (1) an overview of cloning procedures described in this paper, (2) specific Golden Gate reaction protocols and (3) standard primers for cloning and sequencing of plasmids and generation of deletion cassettes by PCR and split-marker PCR. These include standard cassettes for hygromycin B, nourseothricin and phleomycin resistance genes as well as FLP/ FRT-based marker recycling cassettes for hygromycin B and nourseothricin resistance genes. For gene deletion, we provide five different donor vectors for selection marker cassettes. We generated plasmids for C- as well as N-terminal tagging with GFP, mRFP and 3xFLAG. Our vector set contains recognition sites for the commonly used type IIS restriction endonuclease BsaI. In Golden Gate cloning, restriction and ligation occur simultaneously in a one-pot reaction. ![]() Here, we provide Golden Gate vectors for fast and easy cloning of gene fusion as well as gene deletion vectors applicable to diverse fungi. Yet, cloning is a prerequisite for many standard experiments for the functional analysis of genes, including the generation of deletion mutants and the localization of gene products. The cloning of plasmids can be time-consuming or expensive.
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