We (Yao-Guang Liu & Daisuke Shibata) have constructed a TAC-based Arabidopsis thaliana (Columbia accession) genomic DNA library in which each plasmid contains a DNA insert of ~80 kb (Protocol 3). A copy of the library has been donated to the Arabidopsis Biological Resource Center at Ohio State University for basic research purposes. The TAC library has been used at Kazusa DNA Research Institute for the sequencing of chromosomes III and V as part of the A. thaliana genome project. It has become clear during the course of this work that chimeric clones constitute <5% of the total number of clones in the library, and that most clones are relatively stable in E. coli. However, certain chromosomal regions that contain repeated sequences, such as those associated with centromeres, are not maintained stably (H. Kotani, personal communication). The genome sequencing project has resulted in most chromosomal regions being represented almost contiguously by P1 phage genomic clones of ~75 kb (Liu et al., Plant J. 7, 351-358, 1995), and gaps between P1 clones are represented by TAC clones or other BAC clones7. Sixty-one TAC clones have been used for genomic sequencing, and these sequences are available on the Web site of Kazusa DNA Research Institute (http://www.kazusa.or.jp/kaos/).
To check the stability of clones of the TAC library in A. tumefaciens, we purified 50 plasmids from E. coli and introduced them into A. tumefaciens C58C1(MP90) or EHA105. Most TAC plasmids were maintained stably in the A. tumefaciens strains. However, some of the clones were unstable; after transformation of A. tumefaciens by electroporation, several deleted inserts were detected in the transformants. In such instances, colonies that appeared larger than normal on selection agar plates often exhibited deletion of inserts. We therefore recommend checking the integrity of each TAC clone in A. tumefaciens before gene transfer into plants.