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Figure 2 | Virology Journal

Figure 2

From: A general method to eliminate laboratory induced recombinants during massive, parallel sequencing of cDNA library

Figure 2

Effect of RNA concentration on (a) recombination and (b) mutation rates. RNA was extracted from an equal mix of WT and MK virus. SSIII was used to reverse transcribe 160 ng, 1600 ng or 3990 ng as input template. At the highest RNA concentration dNTP’s and primers were reduced 4-fold to mimic the effect of excess RNA template. Resulting cDNA was diluted such that ~2500 copies were input and amplified using an optimised 2-step PCR and reduced (27) cycles to minimise PCR-induced recombination [1]. Recombination and mutation rates are expressed as rate per 1000 nucleotides and the confidence intervals, as estimated via bootstrapping, are shown. Significant differences (at 95%CI) are indicated.

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