We filled gaps in the Pcdh constant region in scaffold_480 and scaffold_160 by PCR using fugu genomic DNA as a template and extended the sequences by identifying and sequencing overlapping cosmid or BAC clones (see Methods).
Here we report crystal structures of the catalytic core of yeast Pol α in unliganded form, bound to an RNA primer/DNA template and extending an RNA primer with deoxynucleotides.
Object-oriented software design patterns are implemented as templates and extended by application software.
Specifically, the 5′-biotinylated primer containing a dU near the primer template junction was hybridized with the adducted templates and extended by the Dpo4 polymerase to generate bypass products.
Once base paired, the reverse transcriptase switches templates and extends into the linker sequences allowing PCR amplification of full-length cDNA using a single primer (the 5'-linker has the same sequences as the linker containing poly-T used for the synthesis of the first strand cDNA).
In the notched screw cap complex, human RFC anchors to the single-stranded region of the P/T junction, covering 12 nucleotides of the template strand, and extends into the double-stranded region, covering 15 and 8 nucleotides within the primer and template strands, respectively (Tsurimoto and Stillman, 1991; Gulbis et al., 1996).
These are annealed to single-stranded dUTP-containing template plasmid and extended with T7 polymerase to create a complementary strand, and the resulting termini are ligated with T4 DNA ligase.
Thermal cycler conditions were 15 min at 50°C for cDNA synthesis, 10 min at 95°C followed by 40 cycles of 15 s at 95°C to denature the DNA, and 45 s at 60°C to anneal and extend the template.
For example, unmethylated extension products were obtained with oligo 28 and oligo 45, which should only bind to and extend methylated templates.
As the DeepLens team stressed during our workshop, even developers who have never worked with machine learning can take the existing templates and easily extend them.
