Computational protocol: Grateloupia tenuis Wang et Luan sp. nov. (Halymeniaceae, Rhodophyta): A New Species from South China Sea Based on Morphological Observation and rbcL Gene Sequences Analysis

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Protocol publication

[…] DNA samples of LNU20092087, LNU20092088, LNU20092089, and LNU20092090 were extracted using the DNeasy Plant Mini Kit (QIAGEN, Valencia, CA, Beijing). The procedures for PCR amplification and sequencing were executed as described previously []. The rbcL gene sequences of 32 extra relevant Halymeniaceae species and two additional related family (Gelidiaceae and Gracilariaceae) species were selected from GenBank for analysis and were involved in the alignment (). Gelidiella ligulata  Dawson and Gracilaria tenuistipitata Chang et Xia were treated as outgroups.The rbcL sequences were aligned and compiled with Clustal X version 1.83 [] for phylogenetic analysis. Phylogenetic tree construction and nucleotide differences analyses were conducted using MEGA 5.0. Maximum parsimony (MP), neighbor joining (NJ), and maximum likelihood (ML) were adopted to construct the phylogenetic tree. The MP analysis used heuristic searches for evaluating tree likelihoods, which was carried out with 1000 replicates, employed random addition sequence of taxa and used tree-bisection-reconnection (TBR) branch swapping []. The NJ analysis used the ratio test for estimation to seek optimal settings and ensure data dependability and was performed with Modeltest version 3.06 []. For ML analysis, a variety of cumulatively complex models of molecular evolution were assessed, as summarized by Litaker et al. [] and Moncalvo et al. []. Bootstrap support values were calculated using 1000 samplings of the dataset [] to estimate statistical reliability for MP, NJ, and ML methods. […]

Pipeline specifications

Software tools Clustal W, MEGA, ModelTest-NG
Application Phylogenetics
Organisms Plectorhinchus orientalis