in subsequent analyses, and they might seriously bias the
results.
33. The list of available Arabidopsis genomes is continuously
increasing through individual laboratory projects. For instance,
additional sequences from African genomes can be downloaded at www.mpipz.mpg.de/hancock/ [18].
34. Exporting vcf or fasta files to specific software packages used
for nucleotide diversity analyses will depend on the format
accepted by each software. Vcf files are directly open, filtered
by SNP frequency, and exported as text files, using multiple
software packages such as TASSEL or DnaSP (see Table 1).
Text files can be easily converted to FASTA files, which can
be imported to common programs for nucleotide diversity
analyses, such as DnaSP or MEGA (see Table 1).
35. Some software programs for the analysis of nucleotide diversity
allow assignment of coding and noncoding regions in your
gene (DnaSP or MEGA; see Table 1). This classification of
gene regions enables separate estimations of nucleotide diversity for synonymous and non-synonymous sites of coding
regions, for noncoding regions, or for noncoding plus synonymous sites (silent diversity).
36. The list of polymorphic sites, their predicted functional effects,
and the missense mutations in your gene can be directly
obtained in the 1001 genomes database (sections 1001 Polymorph, or 1001 Proteomes). Alternatively, this information
can be obtained with software programs for nucleotide diversity analysis (DnaSP or MEGA; see Table 1) after defining
coding and noncoding regions in your gene (see Note 35).
37. High impact mutations detected in a single accession (singletons) should be confirmed by standard DNA sequencing.
Seeds of accessions carrying these mutations can be requested
from Arabidopsis stock centers.
38. NJ trees and PCA can be carried out with TASSEL or MEGA
(see Table 1), which include visualization tools, as well as with
the R package ape developed for analyses of phylogenetics and
evolution [70].
39. The genetic differentiation between groups of accessions
belonging to distinct haplogroups or geographic regions can
be estimated with DnaSP or MEGA (see Table 1). The precise
polymorphisms differentiating such groups can be identified
from sequence alignments sorted according to those groups.
The detection of highly differentiated haplogroups involving
polymorphisms with predicted functional effects might suggest
functionally differentiated alleles in your gene of interest.
However, it cannot be discarded that such haplogroups
106
Carlos Alonso-Blanco et al.
results.
33. The list of available Arabidopsis genomes is continuously
increasing through individual laboratory projects. For instance,
additional sequences from African genomes can be downloaded at www.mpipz.mpg.de/hancock/ [18].
34. Exporting vcf or fasta files to specific software packages used
for nucleotide diversity analyses will depend on the format
accepted by each software. Vcf files are directly open, filtered
by SNP frequency, and exported as text files, using multiple
software packages such as TASSEL or DnaSP (see Table 1).
Text files can be easily converted to FASTA files, which can
be imported to common programs for nucleotide diversity
analyses, such as DnaSP or MEGA (see Table 1).
35. Some software programs for the analysis of nucleotide diversity
allow assignment of coding and noncoding regions in your
gene (DnaSP or MEGA; see Table 1). This classification of
gene regions enables separate estimations of nucleotide diversity for synonymous and non-synonymous sites of coding
regions, for noncoding regions, or for noncoding plus synonymous sites (silent diversity).
36. The list of polymorphic sites, their predicted functional effects,
and the missense mutations in your gene can be directly
obtained in the 1001 genomes database (sections 1001 Polymorph, or 1001 Proteomes). Alternatively, this information
can be obtained with software programs for nucleotide diversity analysis (DnaSP or MEGA; see Table 1) after defining
coding and noncoding regions in your gene (see Note 35).
37. High impact mutations detected in a single accession (singletons) should be confirmed by standard DNA sequencing.
Seeds of accessions carrying these mutations can be requested
from Arabidopsis stock centers.
38. NJ trees and PCA can be carried out with TASSEL or MEGA
(see Table 1), which include visualization tools, as well as with
the R package ape developed for analyses of phylogenetics and
evolution [70].
39. The genetic differentiation between groups of accessions
belonging to distinct haplogroups or geographic regions can
be estimated with DnaSP or MEGA (see Table 1). The precise
polymorphisms differentiating such groups can be identified
from sequence alignments sorted according to those groups.
The detection of highly differentiated haplogroups involving
polymorphisms with predicted functional effects might suggest
functionally differentiated alleles in your gene of interest.
However, it cannot be discarded that such haplogroups
106
Carlos Alonso-Blanco et al.