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Background  Failure of colony PCRs in green microalga  Chlorella vulgaris  is typically attributed to the difficulty in disrupting its notoriously rigid cell walls for releasing the genetic materials and therefore the development of an effective colony PCR procedure in  C. vulgaris  presents a challenge.    Results  Here we identified that colony PCR results were significantly affected by the accumulated lipids rather than the rigid cell walls of  C. vulgaris.  The higher lipids accumulated in  C. vulgaris  negatively affects the effective amplification by DNA polymerase. Based on these findings, we established a simple and extremely effective colony PCR procedure in  C. vulgaris . By simply pipetting/votexing the pellets of  C. vulgaris  in 10 ul of either TE (10 mM Tris/1 mM EDTA) or 0.2% SDS buffer at room temperature, followed by the addition of 10 ul of either hexane or Phenol:Chloroform:Isoamyl Alcohol in the same PCR tube for extraction. The resulting aqueous phase was readily PCR-amplified as genomic DNA templates as demonstrated by successful amplification of the nuclear 18S rRNA and the chloroplast  rbcL  gene. This colony PCR protocol is effective and robust in  C. vulgaris  and also demonstrates its effectiveness in other  Chlorella  species.    Conclusions  The accumulated lipids rather than the rigid cell walls of  C. vulgaris  significantly impede the extraction of genetic materials and subsequently the effective colony PCRs. The finding has the potential to aid the isolation of high-quality total RNAs and mRNAs for transcriptomic studies in addition to the genomic DNA isolation in  Chlorella .

Accumulated lipids rather than the rigid cell walls impede the extraction of genetic materials for effective colony PCRs in Chlorella vulgaris