Open Access Research article

A second generation framework for the analysis of microsatellites in expressed sequence tags and the development of EST-SSR markers for a conifer, Cryptomeria japonica

Saneyoshi Ueno1*, Yoshinari Moriguchi1, Kentaro Uchiyama1, Tokuko Ujino-Ihara1, Norihiro Futamura2, Tetsuya Sakurai3, Kenji Shinohara2 and Yoshihiko Tsumura1

Author Affiliations

1 Department of Forest Genetics, Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki, 305-8687, Japan

2 Department of Molecular and Cell Biology, Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki, 305-8687, Japan

3 Integrated Genome Informatics Research Unit, Plant Science Center, RIKEN Yokohama Institute, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama City, Kanagawa, 230-0045, Japan

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BMC Genomics 2012, 13:136  doi:10.1186/1471-2164-13-136

Published: 16 April 2012

Additional files

Additional file 1:

Table S1. Web links to programs/resources that were used in the present study. Web links were accessed on 19th November 2011.

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Additional file 2:

Table S2. Parameters used in Primer3 program for (a) read2Marker and (b) CMiB pipeline.

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Additional file 3:

cmib. An example shell script for the CMiB pipeline.

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Additional file 4:

Figure S1. Location of C. japonica samples used to screen polymorphisms.

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Additional file 5:

Table S3. EST-SSR markers for C. japonica. Primer sequences are first grouped by pipelines used (read2Marker or CMiB). Forward and reverse primer sequences are listed in upper and lower part of a cell, respectively. Primer sequences include additional bases in 5′ end for fluorescent labelling. Ta: annealing temperature.

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Additional file 6:

Figure S2. Length distribution of reads obtained by (A) Sanger and (B) pyrosequencing method.

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Additional file 7:

Figure S3. (A) Length and (B) depth distribution of Contigs (CjCon1).

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Additional file 8:

Table S4. Assembly statistics from (A) each library, (B) 3′ ESTs and (C) 5′ ESTs.

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Additional file 9:

Figure S4. SSR frequency and density for assemblies in different sequencing direction (3′ EST and 5′ EST) for each library.

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Additional file 10:

Figure S5. Motif frequency of EST-SSRs in CjCon1 and EST contigs for seven species in gene index. The gene indices are assigned as the following abbreviations: AGI; Arabidopsis thaliana, HAGI; Helianthus annuus, NTGI; Nicotiana tabacum, OGI; Oak, OSGI; Oryza sativa, PGI; Pinus and SGI; Picea.

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Additional file 11:

Figure S6. GC percent for CjCon1 and other seven gene indices. The gene indices abbreviations are as follows: AGI; Arabidopsis thaliana, HAGI; Helianthus annuus, NTGI; Nicotiana tabacum, OGI; Oak, OSGI; Oryza sativa, PGI; Pinus and SGI; Picea.

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Additional file 12:

Table S5. Summary of gene ontology annotations for CjCon1. Percentage for each GO was based on the total number of sequences with GO annotation. For CjCon1, 37,387 contigs had GO annotation, while for contigs with SSRs, 1,737 had GO annotation. Cells in yellow indicate GOs that are over-represented in SSR-containing ESTs.

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Additional file 13:

Table S6. Annotation of EST-SSR markers. #N/A indicates no blast hits.

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Additional file 14:

Table S7. Levels of polymorphisms for EST-SSR markers in C. japonica. Twenty-four markers from the top are from read2Marker pipeline, while the rest of the markers are from CMiB pipeline. N: number of individuals genotyped; Na: number of alleles per locus; HO: observed heterozygosity; HE: expected heterozygosity; FIS: fixation index; P value: levels of significance for deviation from Hardy-Weinberg equilibrium; PIC: polymorphism information content.

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