#### Alignment and ML Phylogeny analyses #### #MUSCLE was run in talapas #ML inference IQtree results saved to ML folder #GTR+I+G often overwrite the original iqtree outputs #Gblocks output was NOT determined with the ">" operator, included here just for simplicity. ## DS initial Sequence list with few outgroup ## mafft --auto --inputorder "salps18S.fasta" > "salps18Saligned.fa" ../iqtree -s Seqs_and_Alignments/salps18Saligned.fa -nt AUTO -bb 1000 ## DS Chordata Sequence List with many outgroups included in Gov2011 ## mafft --auto --inputorder "chordata18S.fasta" > "chordata18Saligned.fa" ../iqtree -s Seqs_and_Alignments/chordata18Saligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/chordata18Saligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE ./muscle5.1.linux_intel64 -align chordata18S.fasta -output chordata18S_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/chordata18S_MUSCLE.afa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/chordata18S_MUSCLE.afa-gb -nt AUTO -bb 1000 -m GTR+I+G #with GBLOCKS ./Gblocks chordata18Saligned.fa -t DNA -b5 h > chordata18Saligned.fa-gb ../iqtree -s Seqs_and_Alignments/chordata18Saligned.fa-gb -nt AUTO -bb 1000 #with MUSCLE+GBLOCKS ./Gblocks chordata18S_MUSCLE.afa -t DNA -b5 h > chordata18S_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/chordata18S_MUSCLE.afa-gb -nt AUTO -bb 1000 ## Original Gov2011 accessions without AY903925.1 Halocynthia igaboja ## mafft --auto --inputorder "Govindarajan2011_18S.fasta" > "Govindarajan2011_18Saligned.fa" ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18Saligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/Govindarajan2011_GTRIG.fa -nt AUTO -bb 1000 -m GTR+I+G #Removing the alleged typoed Uncultured bacterium accession HQ015280.1 (DELETED files) mafft --auto --inputorder "Govindarajan2011-UncBac_18S.fasta" > "Govindarajan2011-UncBac_18Saligned.fa" ../iqtree -s Seqs_and_Alignments/Govindarajan2011-UncBac_18Saligned.fa -nt AUTO -bb 1000 ## Original Gov2011 accessions with AY903925.1 Halocynthia igaboja ## mafft --auto --inputorder "Govindarajan2011_18S+Higaboja.fasta" > "Govindarajan2011_18S+Higaboja_aligned.fa" ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_aligned.fa -nt AUTO -bb 1000 #with GBLOCKS ./Gblocks Govindarajan2011_18S+Higaboja_aligned.fa -t DNA -b5 h > Govindarajan2011_18S+Higaboja_GBLOCK.fa-gb ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_GBLOCK.fa-gb -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_GBLOCK.fa-gb -nt AUTO -bb 1000 -m GTR+I+G #models limited to raxml selection ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_GBLOCK.fa-gb -nt AUTO -bb 1000 -m GTR+I+G -mset raxml #with specific pinv and alpha ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_GBLOCK.fa-gb -nt AUTO -bb 1000 -m GTR+I+G -a 0.4861 -i 0.3784 #with MUSCLE ./muscle5.1.linux_intel64 -align Govindarajan2011_18S+Higaboja.fasta -output Govindarajan2011_18S+Higaboja_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_MUSCLE.afa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_MUSCLE.afa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE+GBLOCKS ./Gblocks Govindarajan2011_18S+Higaboja_MUSCLE.afa -t DNA -b5 h > Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_MUSCLE.afa-gb -nt AUTO -bb 1000 #with GTR+I+G #with specific pinv and alpha ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja_MUSCLE.afa-gb -nt AUTO -bb 1000 -m GTR+I+G -a 0.4861 -i 0.3784 ## UNION between DS Chordata Sequence List and Gov2011 including AY903925.1 H. igaboja ## #All Govindarajan sequences + AJ250778 Ciona intestinalis and AB921975 Molgula manhattensis mafft --auto --inputorder "GovindarajanUNIONChordata.fasta" > "GovindarajanUNIONChordata_aligned.fa" ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE ./muscle5.1.linux_intel64 -align GovindarajanUNIONChordata.fasta -output GovindarajanUNIONChordata_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_MUSCLE.afa -nt AUTO -bb 1000 #with GBLOCKS ./Gblocks GovindarajanUNIONChordata_MUSCLE -t DNA -b5 h > GovindarajanUNIONChordata_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_aligned.fa-gb -nt AUTO -bb 1000 #with MUSCLE+GBLOCKS ./Gblocks GovindarajanUNIONChordata_aligned.fa -t DNA -b5 h > GovindarajanUNIONChordata_aligned.fa-gb ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_MUSCLE.afa-gb -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovindarajanUNIONChordata_MUSCLE.afa-gb -nt AUTO -bb 1000 -m GTR+I+G ## INTERSECTION between DS Chordata Sequence List and Gov2011 including AY903925.1 H. igaboja ## mafft --auto --inputorder "chordata18S-Mman-Cint.fasta" > "chordata18S-Mman-Cint_aligned.fa" ../iqtree -s Seqs_and_Alignments/chordata18S-Mman-Cint_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/chordata18S-Mman-Cint_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE ./muscle5.1.linux_intel64 -align chordata18S-Mman-Cint.fasta -output chordata18S-Mman-Cint_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/chordata18S-Mman-Cint_MUSCLE.afa -nt AUTO -bb 1000 ## Gov2011 including AY903925.1 H. igaboja + AB921975 Molgula manhattensis ## #Labeled as DS-Union-Gov11 minus C. intestinalis mafft --auto --inputorder "GovUNIONCho-AJ250778Cint.fasta" > "GovUNIONCho-AJ250778Cint_aligned.fa" ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AJ250778Cint_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AJ250778Cint_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE ./muscle5.1.linux_intel64 -align GovUNIONCho-AJ250778Cint.fasta -output GovUNIONCho-AJ250778Cint_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AJ250778Cint_MUSCLE.afa -bb 1000 -nt AUTO #with MUSCLE+GBLOCKS ./Gblocks GovUNIONCho-AJ250778Cint_MUSCLE.afa -t DNA -b5 h > GovUNIONCho-AJ250778Cint_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AJ250778Cint_MUSCLE.afa-gb -nt AUTO -bb 1000 ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis ## #Labeled as DS-Union-Gov11 minus M. manhattensis mafft --auto --inputorder "GovUNIONCho-AB921975Mman.fasta" > "GovUNIONCho-AB921975Mman_aligned.fa" ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with MUSCLE ./muscle5.1.linux_intel64 -align GovUNIONCho-AB921975Mman.fasta -output GovUNIONCho-AB921975Mman_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_MUSCLE.afa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_MUSCLE.afa -nt AUTO -bb 1000 -m GTR+I+G #with GBLOCKS ./Gblocks GovUNIONCho-AB921975Mman_aligned.fa -t DNA -b5 h > GovUNIONCho-AB921975Mman_aligned.fa-gb ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_aligned.fa-gb -nt AUTO -bb 1000 #with MUSCLE+GBLOCKS ./Gblocks GovUNIONCho-AB921975Mman_MUSCLE.afa -t DNA -b5 h > GovUNIONCho-AB921975Mman_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_MUSCLE.afa-gb -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_MUSCLE.afa-gb -nt AUTO -bb 1000 -m GTR+I+G ## Original Gov2011 accessions with AY903925.1 Halocynthia igaboja and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti ## mafft --auto --inputorder "Govindarajan2011_18S+Higaboja+news.fasta" > "Govindarajan2011_18S+Higaboja+news_aligned.fa" ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja+news_aligned.fa -nt AUTO -bb 1000 #with MUSCLE ./muscle5.1.linux_intel64 -align Govindarajan2011_18S+Higaboja+news.fasta -output Govindarajan2011_18S+Higaboja+news_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja+news_MUSCLE.afa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/Govindarajan2011_18S+Higaboja+news_MUSCLE.afa -nt AUTO -bb 1000 -m GTR+I+G #_MUSCLE-gtrig #with GBLOCKS ./Gblocks Govindarajan2011_18S+Higaboja+news_aligned.fa -t DNA -b5 h > Govindarajan2011_18S+Higaboja+news.fa-gb #really bad % sites ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti ## #Labeled as DS-Union-Gov11 minus M. manhattensis + news mafft --auto --inputorder "GovUNIONCho-AB921975Mman+news.fasta" > "GovUNIONCho-AB921975Mman+news_aligned.fa" ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman+news_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #-gtrig #with MUSCLE ./muscle5.1.linux_intel64 -align GovUNIONCho-AB921975Mman+news.fasta -output GovUNIONCho-AB921975Mman+news_MUSCLE.afa ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman+news_MUSCLE.afa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman+news_MUSCLE.afa -nt AUTO -bb 1000 -m GTR+I+G #_MUSCLE-gtrig. #with GBLOCKS ./Gblocks GovUNIONCho-AB921975Mman+news_aligned.fa -t DNA -b5 h > GovUNIONCho-AB921975Mman+news_aligned.fa-gb ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman+news_aligned.fa-gb -nt AUTO -bb 1000 #with MUSCLE+GBLOCKS ./Gblocks GovUNIONCho-AB921975Mman+news_MUSCLE.afa -t DNA -b5 h > GovUNIONCho-AB921975Mman+news_MUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/GovUNIONCho-AB921975Mman+news_MUSCLE.afa-gb -nt AUTO -bb 1000 ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti + the SANGER SEQUENCED ## #Labeled as GUC-Mm+N+Sanger mafft --auto --inputorder "Sanger/GUC-Mm+N+Sanger.fa" > "GUC-Mm+N+Sanger_aligned.fa" ../iqtree -s GUC-Mm+N+Sanger_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s GUC-Mm+N+Sanger_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with GBLOCKS didntwork ###./Gblocks GUC-Mm+N+Sanger_aligned.fa -t DNA -b5 h > GUC-Mm+N+Sanger_aligned.fa-gb ### #with MUSCLE ../muscle5.1.macos_arm64 -align Sanger/GUC-Mm+N+Sanger.fa -output GUC-Mm+N+Sanger_MUSCLE.afa ../iqtree -s GUC-Mm+N+Sanger_MUSCLE.afa -nt AUTO -bb 1000 ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti + the SANGER SEQUENCED SECOND BATCH ## #Labeled as GUC-Mm+N+Sanger2 mafft --auto --inputorder "Sanger/GUC-Mm+N+Sanger2.fa" > "GUC-Mm+N+Sanger2_aligned.fa" ../iqtree -s GUC-Mm+N+Sanger2_aligned.fa -nt AUTO -bb 1000 #with GTR+I+G ../iqtree -s GUC-Mm+N+Sanger2_aligned.fa -nt AUTO -bb 1000 -m GTR+I+G #with GBLOCKS ./Gblocks Seqs_and_Alignments/GUC-Mm+N+Sanger2_aligned.fa -t DNA -b5 h > Seqs_and_Alignments/GUC-Mm+N+Sanger2_aligned.fa-gb ../iqtree -s Seqs_and_Alignments/GUC-Mm+N+Sanger2_aligned.fa-gb -nt AUTO -bb 1000 #with MUSCLE ../muscle5.1.macos_arm64 -align Sanger/GUC-Mm+N+Sanger2.fa -output GUC-Mm+N+Sanger2_MUSCLE.afa ../iqtree -s GUC-Mm+N+Sanger2_MUSCLE.afa -nt AUTO -bb 1000 #with GBLOCKS ./Gblocks GUC-Mm+N+Sanger2_MUSCLE.afa -t DNA -b5 h > GUC-Mm+N+Sanger2_aMUSCLE.afa-gb ../iqtree -s Seqs_and_Alignments/GUC-Mm+N+Sanger2_MUSCLE.afa-gb -nt AUTO -bb 1000 ###### ### BAYESIAN Analyses in RevBayes #### #Topology ../rb topology.Rev #Ultrametric molecular clock branch length estimation ../rb mcmc_TimeTree_salps.Rev #Directories: ## DS Chordata Sequence List with many outgroups included in Gov2011 aligned## #with MAFFT TOPOLOGY_Chordata_output #timetree TIMETREE_Chordata_output ## Original Gov2011 accessions with AY903925.1 Halocynthia igaboja ## #with MAFFT TOPOLOGY_Gov2011+Higaboja_output ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis ## #with MUSCLE TOPOLOGY_Gov2011+Higa+Cint_MUSCLE_output #timetree TIMETREE_Gov2011+Higa+Cint_MUSCLE_output #with MUSCLE+GBLOCKS TOPOLOGY_Gov2011+Higa+Cint_MUSCLEGB_output ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti ## #with MUSCLE TOPOLOGY_Gov2011+HigaCint+New_MUSCLE_output #with MAFFT TOPOLOGY_Gov2011+HigaCint+New_MAFFT_output ## Gov2011 including AY903925.1 H. igaboja + AJ250778 Ciona intestinalis and the NEW SEQUENCES for salps B. lacromae, T. longicauda, S. younti + the SANGER SEQUENCED ## #Labeled as GUC-Mm+N+Sanger #with MUSCLE TOPOLOGY_GUC-Mm+N+Sanger_MUSCLE_output #timetree TIMETREE_Gov2011+Higa+Cint_MUSCLE_output