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This template/assign strategy is flexible, as templates can be produced from single samples to establish a master template; combinations of samples to produce a consensus template; or stochastic subsets of cells to optimize run times and the sensitivity of detection of small clusters.
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Quantification for each primer pair and cDNA template combination was performed in triplicate for each of the two biological replicates at every time point.
During the annealing step the most complementary primer-template combination gives rise to a primary product which is then amplified further in a secondary PCR reaction.
Each primer-template combination was run with three technical replicates and three biological replicates.
The efficacy of extension from these templates was measured by determining the kinetic parameters for incorporation of the next correct nucleotide (dG) to each primer-template combination.
This being the case, all three of our predictors which implemented this portion of the pipeline incorporated the multi-template combination algorithm (Cheng, 2008) (Table 1).
Our multi-level combination pipeline (Fig. 1) for protein structure prediction is generally comprised of five steps: (i) template identification and ranking, (ii) multi-template combination, (iii) model generation, (iv) model evaluation and (v) model combination and refinement.
Sir John's multidrug approach remains the standard for treating tuberculosis and has provided the template for combination therapies to combat grave diseases like cancer and AIDS.
3 µl of the cDNA template in combination with 0.2 mM dNTP mix, 2 mM MgSO4, 0.25 µM each primer, and 1 U Platinum Taq high fidelity polymerase (Invitrogen) was used in a standard PCR reaction.
The new template in combination with the refined machine-learned score function was used to identify amino acid substitutions that either destabilized interactions with d-Thy or resulted in structural changes favoring bound l-Thy in the phosphoryl-acceptor site.
Each of the CPDL variants was constructed by overlap extension PCR using AtFatB-par as template in combination with the primers listed in Table 1 then cloned into the pBC expression plasmid.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com