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*演習2 [#p1e5cf33]
**Major evolutionary transitions in ant agriculture [#ma7c1896]
-Ted R. Schultz and Sean G. Brady. 2008~
[[PNAS. 105(14): 5435-5440.>http://www.pnas.org/content/105/14/5435.full]]


***Data [#l57a712c]
-2,459 aligned nucleotide sites from the coding regions of four nuclear genes:
--elongation factor 1-F1 (EF1-F1) (1,075 bp)
--elongation factor 1-F2 (EF1-F2) (517 bp)
--wingless (409 bp)
--long-wavelength rhodopsin (opsin) (458 bp)
-All data in this study represent protein-coding (exon) sequences~
intervening introns in opsin and EF1F1 were not used because they could not be aligned confidently. 
-Sample: 65 attine taxa and 26 nonattine outgroups.~
Primers used for PCR amplification and sequencing are found in supporting information (SI) Table S1.
-Of the total 2,459 included nucleotide positions from all genes, 952 were variable and 847 parsimony informative. Sequences are deposited in GenBank; taxa and accession numbers are listed in Table S2.

***Phylogenetic Analyses [#u92e7045]
-(i) Maximum parsimony (MP) analyses
--PAUP* v4.0b10
---heuristic searches with tree bisection.reconnection (TBR) and 1,000 random-taxon-addition replicates. ~
Analyses identified 12 most-parsimonious trees (MPTs) of length  4,383, CI  0.270, RI  0.704. Successive-approximations-weighting analyses identified a single tree, one of the MPTs.
---Nonparametric bootstrap analyses used TBR branch-swapping and consisted of 1,000 pseudoreplicates, with 10 random-taxon-addition replicates per pseudoreplicate. 

-(ii) Maximum likelihood (ML)
-- ModelTest v3.06~
The data and the MPT identified by weighting were evaluated under the Akaike information criterion (AIC) as calculated in, 
---identifying the GTR model of evolution. 
--GARLI v0.951 using the GTR model (with six  rate categories), with a heuristiclosuccessiveapproximationsg likelihood of 24,868.84927. 
---Nonparametric bootstrap analyses consisted of 500 pseudoreplicates in GARLI under the same conditions as the ML search.
---A subsequent  search in PAUP* using the most likely tree identified by the GARLI searches as the starting tree and employing TBR branch-swapping and the GTRI model (with six  rate categories) resulted in exactly the same topology and likelihood score. 

-(iii) Bayesian nucleotide-model Markov Chain Monte Carlo (MCMC):~
MrBayes v3.1.2 (59). 
--Burn-in and run convergence were assessed by comparing the mean and variance of log likelihoods, both by eye and by using the program
---Tracer v1.3
---MrBayes  e e.stat f f output file
---MrBayes bthe split frequencies diagnostic.
--Eight character partitions for nucleotide-model analyses:
---four partitions consisting of the combined first and second codon positions for each of the four genes
---four partitions consisting of the third codon position for each of the four genes.
---based on ModelTest results~
the wingless third-position - GTR model~
opsin and EF1F2 third positions - separately assigned the HKYI model~
all other character partitions - separately assigned the GTRI model
-(iv) Bayesian codon-model MCMC~

**Phylogenetic Mapping of Agricultural Systems. [#m346c506]
//-Terminal taxa were assigned states for a single six-state character representing the four attine agricultural systems and leaf-cutter agriculture (i.e., no agriculture, lower agriculture, yeast agriculture, higher agriculture, leaf-cutter agriculture, coral-fungus agriculture).
//-Five species (Myrmicocrypta n. sp. Brazil, Mycetagroicus triangularis, Cyphomyrmex n. sp., Cyphomyrmex morschi, Trachymyrmex irmgardae, and Pseudoatta n. sp.) received  e eunknown f f (i.e.,  e e? f f) state assignments, and @Trachymyrmex papulatus received a  e elower agriculture f f state assignment based on a single garden collection from Argentina (a second colony from the same locality cultivated a typical higher attine garden). 
//-Character evolution was optimized onto the Bayesian codon-model consensus tree (with branch lengths) under both parsimony using MacClade and maximum likelihood using the StochChar module provided in the Mesquite package.
//-Under parsimony, ancestral-state optimizations were unambiguous. 
//-Under the Markov k-state 1-parameter model,  the likelihood that each agricultural system arose in the most recent common ancestor of the corresponding ant clade was, as a proportion of the total probability ( 1.0) distributed across the six character states, 0.9831 for lower agriculture, 0.9995 for yeast agriculture, 0.9905 for higher agriculture, 0.9924 for leaf-cutter agriculture, and 0.9998 for coral-fungus agriculture.

**Divergence Dating [#gae0451f]
//-We inferred divergence dates using both semiparametric and Bayesian relaxed clock methods.
//-The first method used was the semiparametric penalized likelihood approach implemented in r8s v1.7 (64, 65). 
//--Branch lengths were first estimated on the ML topology using PAUP* under a GTRI model. 
//--The Pogonomyrmex and two Myrmica species were used to root the tree during branch length estimation and were subsequently removed from all dating analyses. 
//--Thus, the root of the tree for all dating analyses represents the origin of the  e ecore myrmicines, f f a well supported clade established by previous work (33). 
//--Smoothing parameters were estimated by using the cross-validation feature in r8s. 
//--Confidence intervals were calculated by using 100 nonparametric bootstrap replicates of the dataset generated by Mesquite, followed by reestimation of branch lengths and divergence times for each replicate.
//-We calibrated three nodes with minimum-age constraints using attine Dominican amber fossils. 
//^^These fossils are 
//---(i) Apterostigma electropilosum, a member of the A. pilosum group 
//---(ii) Cyphomyrmex maya and Cyphomyrmex taino, both members of the C. rimosus group 
//---(iii) Trachymyrmex primaevus, a fossil of uncertain placement within the genus (but see below).