Table 1.
The nine phytoplankton species used in this study.
Table 2.
The sequences and annealing temperatures (Ta’s) of the specific primers of ITS used in qPCR.
Fig 1.
Comparison between bead-beating method and non-bead-beating method in estimated cellular DNA content.
Shown are averages, with error bars indicating standard deviations (n = 15).
Table 3.
The comparisons of DNA yields and ITS copies between bead-beating and non-bead-beating methods for the 9 phytoplanktonic species (t-test).
Fig 2.
Gel electrophoretic result of extracted DNA indicating similar DNA integrity between bead-beating (right of lane M) and non-bead-beating (left of lane M) methods.
Lanes 1, 10: A. fundyense; 2,11: K. mikimotoi; 3, 12: P. donghaiense; 4,13: S. costatum; 5,14: C. muelleri; 6,15: T. weissflogii; 7,16: I. galbana; 8,17: Chlorella sp.; 9,18: H. akashiwo. M is GeneRuler 1kb DNA Ladder (Thermo Scientific, USA) with the largest size of 10 kb.
Table 4.
Comparison of DNA purities resulting from the two DNA extraction methods indicated by A260/A280 and A260/A230 ratios (mean ± standard deviation; n = 15).
Fig 3.
The amplification of the whole ITS region using primers 18ScomF-3end and com28SR2 with bead-beating extracted DNA as template.
Different sizes of the amplicons reflected different lengths of the ITS region among different species. Lanes 1–9, A. fundyense, K. mikimotoi, P. donghaiense, S. costatum, C. muelleri, T. weissflogii, I. galbana, Chlorella sp., H. akashiwo, respectively. Lane M, DL2000 DNA Marker (Takara Bio, Japan).
Fig 4.
Comparison of bead-beating with non-bead-beating methods in terms of DNA contents and ITS copy number.
Shown are averages, with error bars indicating standard deviations (n = 15). No significant difference was found between the ratios of DNA contents and those of ITS copies in any of the species examined (p > 0.05, t-test).
Table 5.
ITS copies per cell calculated from qPCR results obtained from DNA templates prepared using the two DNA extraction methods (mean ± standard deviation; n = 15).
Table 6.
Genome sizes (Gbpa) of the 7 phytoplankton species measured using flow cytometric and DNA extraction methods (mean ± standard deviation).
Table 7.
The number of cells and their ratios of the three algal species added into the field sample and those estimated using qPCR.
Fig 5.
The phylogenetic affiliations (A) and taxa composition (B) of the field samples obtained from the retrieved rDNA clone library.
In (A), values at the nodes are bootstrap values derived from Neighbor-joining and Maximum Likelihood methods (NJ/ML); only those larger than 50/0.50 are shown. The branches consisting of only our retrieved clones (OTUs) (except ①, ②, ③ and ④) were collapsed. In (B), 1–9 refer to Archaea (1.1%), Amoebozoa (4.97%), Chlorophyta (21.55%), Ciliophora (7.18%), Bacillariophyta (37.02%), Dinophyta (7.73%), Fungi (5.52%), Metazoa (jellyfish and copepods; 12.15%), undefined Stramenopiles and Eukaryota (2.76%).
Table 8.
The taxa composition revealed by Lowest Common Ancestors (LCA) and corresponding OTU numbers.