Construction of a binary BAC library for an apomictic monosomic addition line of Beta corolliflora in sugar beet and identification of the clones derived from the alien chromosome

• Published in: Theoretical and applied genetics 2004-05, Vol.108 (7), p.1420-1425
• Main Authors: Fang, X, Gu, S, Xu, Z, Chen, F, Guo, D, Zhang, H.B, Wu, N
• Format: Article
• Language: English
• Subjects: Agriculture - methods; bacterial artificial chromosome clones; bacterial artificial chromosomes; Beta; Beta corolliflora; Beta vulgaris; Beta vulgaris - genetics; Beta vulgaris subsp. vulgaris; Blotting, Southern; chromosome addition; Chromosomes, Artificial, Bacterial - genetics; Chromosomes, Plant - genetics; Cloning; Crosses, Genetic; Deoxyribonucleic acid; DNA; DNA libraries; DNA sequencing; DNA, Complementary - genetics; Electrophoresis, Agar Gel; Ethylenediaminetetraacetic acid; Gene Library; genetic transformation; Genetics; Genomics; interspecific hybridization; Interspersed Repetitive Sequences - genetics; Libraries; nucleic acid hybridization; Nucleotide sequencing; repetitive sequences; Reproduction - genetics; sugar beet; Tetracycline; Tetracyclines; wild relatives
• ISSN: 0040-5752; 1432-2242
• DOI: 10.1007/s00122-003-1566-8

A plant-transformation-competent binary BAC library was constructed from the genomic DNA of the chromosome 9 monosomic addition line of Beta corolliflora Zoss. in sugar beet (B. vulgaris. L). This monosomic addition line (designated M14) is characterized by diplosporic reproduction caused by the alien chromosome carrying the gene(s) responsible for diplospory. The library consists of 49,920 clones with an average insert size of 127 kb, representing approximately 7.5 haploid genome equivalents and providing a greater than 99% probability of isolating a single-copy DNA sequence from the library. To develop the scaffold of a physical map for the alien chromosome, B. corolliflora genome-specific dispersed repetitive DNA sequences were used as probes to isolate BAC clones derived from the alien chromosome in the library. A total of 2,365 positive clones were obtained and arrayed into a sublibrary specific for B. corolliflora chromosome 9 (designated bcBAC-IX). The bcBAC-IX sublibrary was further screened with a subtractive cDNA pool generated from the ovules of M14 and the floral buds of B. vulgaris by the suppression subtractive hybridization method. One hundred and three positive binary BACs were obtained, which potentially contain the genes of the alien chromosome specifically expressed during the ovule and embryo development of M14, and may be associated with apomictic reproduction. Thus, these binary BAC clones will be useful for identification of the genes for apomixis by genetic transformation.