Developing expressed sequence tag libraries and the discovery of simple sequence repeat markers for two species of raspberry (Rubus L.).
BACKGROUND
Due to a relatively high level of codominant inheritance and transferability within and among taxonomic groups, simple sequence repeat (SSR) markers are important elements in comparative mapping and delineation of genomic regions associated with traits of economic importance. Expressed sequence tags (ESTs) are a source of SSRs that can be used to develop markers to facilitate plant breeding and for more basic research across genera and higher plant orders.


METHODS
Leaf and meristem tissue from 'Heritage' red raspberry (Rubus idaeus) and 'Bristol' black raspberry (R. occidentalis) were utilized for RNA extraction. After conversion to cDNA and library construction, ESTs were sequenced, quality verified, assembled and scanned for SSRs.  Primers flanking the SSRs were designed and a subset tested for amplification, polymorphism and transferability across species. ESTs containing SSRs were functionally annotated using the GenBank non-redundant (nr) database and further classified using the gene ontology database.


RESULTS
To accelerate development of EST-SSRs in the genus Rubus (Rosaceae), 1149 and 2358 cDNA sequences were generated from red raspberry and black raspberry, respectively. The cDNA sequences were screened using rigorous filtering criteria which resulted in the identification of 121 and 257 SSR loci for red and black raspberry, respectively. Primers were designed from the surrounding sequences resulting in 131 and 288 primer pairs, respectively, as some sequences contained more than one SSR locus. Sequence analysis revealed that the SSR-containing genes span a diversity of functions and share more sequence identity with strawberry genes than with other Rosaceous species.


CONCLUSIONS
This resource of Rubus-specific, gene-derived markers will facilitate the construction of linkage maps composed of transferable markers for studying and manipulating important traits in this economically important genus.
USDA-ARS, National Clonal Germplasm Repository, 33447 Peoria Road, Corvallis, OR, 97333-2521, USA. Jill.Bushakra@ars.usda.gov.
Jill M
JM
Bushakra
USDA-ARS, Beltsville Agricultural Research Center, Genetic Improvement of Fruits and Vegetables Lab, Bldg. 010A, BARC-West, 10300 Baltimore Ave., Beltsville, MD, 20705-2350, USA. Kim.Lewers@ars.usda.gov.
Kim S
KS
Lewers
Department of Entomology and Plant Pathology, University of Tennessee, 2505 EJ Chapman Drive, 370 PBB, Knoxville, TN, 37996, USA. mstaton1@utk.edu.
Margaret E
ME
Staton
Genomics & Computational Biology Laboratory, Biosystems Research Complex, Clemson University, 51 New Cherry St., 304, Clemson, SC, 29634, USA. tzhebe@clemson.edu.
Tetyana
T
Zhebentyayeva
Genomics & Computational Biology Laboratory, Biosystems Research Complex, Clemson University, 51 New Cherry St., 304, Clemson, SC, 29634, USA. Saski@clemson.edu.
Christopher A
CA
Saski
D030541:Databases, Genetic; D020224:Expressed Sequence Tags; D015723:Gene Library; D063990:Gene Ontology; D005819:Genetic Markers; D018895:Microsatellite Repeats / Q000235:genetics*; D065856:Rubus / Q000235:genetics*; D013045:Species Specificity
D005819:Genetic Markers