Sequence-tagged microsatellite site markers for chickpea (Cicer arietinum L.)

Two small-insert genomic libraries of chickpea (Cicer arietinum L.) were screened with a set of microsatellite-specific oligonucleotide probes. A total of 121 positive clones were identified among 13,000 plated colonies. Thirty-nine clones were recognized by (TAA)5, 26 by (GA)8, 18 by (GT)8, 27 by a pool of AT-rich trinucleotide repeats [(CAA)5, (CAT)5, and (GAA)5], and 11 by a pool of GC-rich trinucleotides [(TCC)5, (CAC)5, (CAG)5, and (CGA)5]. Of 53 clones selected for sequencing, 43 carried a microsatellite. Flanking primer pairs were designed for 28 loci, and used on a small test-set comprising one C. reticulatum and four C. arietinum accessions. Separation of the PCR products on agarose or polyacrylamide gels revealed single bands of the expected size with 22 of the primer pairs. Sixteen of these "Cicer arietinum sequence-tagged microsatellite site" (CaSTMS) markers were polymorphic at an intraspecific level, detecting 2-4 alleles within the four accessions examined. Primer pairs CaSTMS10 and CaSTMS15 revealed 25 and 16 alleles among 63 C. arietinum accessions from different geographic locations, reflecting gene diversity values of 0.937 and 0.922, respectively. Mendelian inheritance of CaSTMS markers was demonstrated using a set of recombinant inbred lines and their parents.