plasmid based mixes  (Addgene inc)

Journal: Nucleic acids research

Article Title: Splicing factor Prp18p promotes genome-wide fidelity of consensus 3'-splice sites.

doi: 10.1093/nar/gkad968

Figure Lengend Snippet: Figure 5. Slu7p is necessary to recruit Prp18p but not sufficient to fulfill its 3 ′ SS fidelity functions. ( A ) Spot dilution growth assay for wild-type and prp18 upf1 strain complemented with Yep24 plasmids expressing PRP18 or o v ere xpressing SLU7 or empty Yep24 (vector). ( B ) RT-PCR analysis of NYV1 alternative 3 ′ SS splicing in WT, upf1 , prp18 and prp18 upf1 strains transformed with a vector (–) or a plasmid overexpressing SLU7 (+). ( C ) R T-PCR analy sis of NYV1 and MUD1 alternativ e splicing in v arious slu7 mutants. All strains are in a upf1 back ground. ( D ) Quantification of the ratio of usage of the major alternative 3 ′ SS of NYV1 and MUD1 relative to the main 3 ′ SS in the various slu7 mutants. Legends as in Figure 4 .

Article Snippet: Construction of the PRP18 / pUG35 and PRP18 / YEp24 base plasmids was accomplished using Gibson assembly ( 23 ) ( New England Biolabs #E2611 ) .

Techniques: Growth Assay, Expressing, Plasmid Preparation, Reverse Transcription Polymerase Chain Reaction, Transformation Assay