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Image Search Results

Journal: Nucleic Acids Research
Article Title: Advancing uracil-excision based cloning towards an ideal technique for cloning PCR fragments
doi: 10.1093/nar/gkl635
Figure Lengend Snippet: Overview of the USER cloning technique. A PacI cassette containing USER vector (upper left corner) is digested with PacI and Nt.BbvCI to generate 8 nt single-stranded 3′ overhangs. A PCR fragment amplified with compatible uracil-containing primers by the PfuTurbo ® C x Hotstart DNA polymerase is mixed with USER™ enzyme mix (removing uracils, pink) and the linearized vector. The mixture is incubated 20 min at 37°C and 20 min at 25°C, and the hybridized product is ready to be transformed into E.coli without prior ligation. Nt.BbvCI recognition sites are marked in tan, PacI recognition sites are marked in light blue. Yellow and green mark single base differences between the generated 3′ overhangs, which are responsible for the directional insertion of the PCR fragment.
Article Snippet: PCR conditions PCR with the following DNA polymerases: HotMaster™ Taq DNA Polymerase (Eppendorf), Platinum® Taq DNA Polymerase (Invitrogen), Pwo DNA Polymerase (Roche), Phusion™ DNA Polymerase (Finnzymes), and
Techniques: Clone Assay, Plasmid Preparation, Polymerase Chain Reaction, Amplification, Incubation, Transformation Assay, Ligation, Generated

Journal: Nucleic Acids Research
Article Title: Advancing uracil-excision based cloning towards an ideal technique for cloning PCR fragments
doi: 10.1093/nar/gkl635
Figure Lengend Snippet: Characteristics and applications of the USER technique. ( a ) A comparison of the ability of DNA polymerases to amplify DNA fragments using uracil-free (-dU) and uracil-containing primers (+dU). Hm Taq: HotMaster™ Taq DNA Polymerase, Taq: Platinum ® Taq DNA Polymerase, Pwo: Pwo DNA Polymerase, Phusion: Phusion™ DNA Polymerase, PfuCx: PfuTurbo ® C x Hotstart DNA polymerase. ( b ) Functional expression of A.thaliana glucose transporter AtSTP1 in X.laevis oocytes from the Xenopus -specific USER compatible vector, pNB1u. H 2 O: control oocytes injected with water; STP1: oocytes injected with AtSTP1 cRNA. ( c ) Functional expression of the cyano fluorescence protein in leaf and root of A.thaliana from the CaMV 35S promoter using the USER compatible pCAMBIA230035Su vector (35S:CFP). Control: empty pCAMBIA230035Su.
Article Snippet: PCR conditions PCR with the following DNA polymerases: HotMaster™ Taq DNA Polymerase (Eppendorf), Platinum® Taq DNA Polymerase (Invitrogen), Pwo DNA Polymerase (Roche), Phusion™ DNA Polymerase (Finnzymes), and
Techniques: Functional Assay, Expressing, Plasmid Preparation, Injection, Fluorescence