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系统发育数据分析步骤求助 已有1人参与
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Sequences were aligned with CLUSTALW (Thompson et al.1994) and the alignment was manually refined using Genedoc (Nicholas et al.1997). The aligned matrix was subjected to a neighbour-joining (NJ) analysis using the Kimura two parameter distance estimates (Kimura 1980). Constant sites were subsequently removed from the alignments. Then the matrix was first subjected to a maximum-likelihood (ML) analysis, searching for the appropriate evolutionary model using Modeltest (Posada and Crandall 1998), and then inputting the appropriate model(s) obtained from the likelihood ratio tests and the Akaike information criterion (AIC), into the ML analysis. All analyses were carried out using (Swofford 1998). Because some sequences from the same species were diverged into different clades, we compared the GC content, the sequence length, and the nucleotide difference among them. Based on the methods of Mayol and Rosselló (2001) we identified the rDNA sequences that appeared to represent paralogous loci. These sequences were excluded in subsequent phylogenetic analysis. The consensus sequence for each group was created with BioEdit (Hall 1999) and aligned with CLUSTAL W (Thompson et al.1994) to show the major difference of the types. The phylogenetic analyses based on the reduced sequence matrix were conducted using parsimony and distance methods with PAUP* 4.0 (Swofford 1998). We first carried out a NJ analysis using the Kimura two parameter distance estimates (Kimura 1980). Maximum parsimony (MP) analyses were performed by heuristic search, tree bisection reconnection (TBR) branch swapping, and RANDOM stepwise addition with 100 replicates. To find the evolutionary model that fits best for given data, we subjected the alignment of the sequences to Modeltest (Posada and Crandall 1998) v3.7. The parameters of the best fit model were then used as input for a ML analysis. Topological robustness was assessed by bootstrap analysis with 500 replicates (Felsenstein 1985). In all analyses the functional outgroup (FOG) (Watrous and Wheeler 1981) used was P. heteroclada. 求大神解决几个问题：第一：Genedoc里要矫正到什么程度就可以了？第二：The aligned matrix was subjected to a neighbour-joining (NJ) analysis using the Kimura two parameter distance estimates。这句话是在CLUSTALW 里进行操作，还是在PAUP 4.0里？第三：Constant sites were subsequently removed from the alignments.为什么要删除Constant sites 后再进行ML分析？第四： inputting the appropriate model(s) obtained from the likelihood ratio tests and the Akaike information criterion (AIC), into the ML analysis.找到最适模型后怎么导入到ML分析中？第五：MP树是什么时候构建？万能的大神快来帮帮我吧，已经卡在这个点上半个多月了。。。 |
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