Mitotic stability of a coding DNA sequence-free version of Leishmania major chromosome 1 generated by targeted chromosome fragmentation

Gene. 2002 May 1;289(1-2):151-9. doi: 10.1016/s0378-1119(02)00506-1.


The deletion of a 260-kb segment containing all the coding DNA sequences (CDS) of chromosome 1 of Leishmania major Friedlin strain was performed through homologous recombination during a transfection experiment. This allowed the selection of a mutant clone containing a linear extra chromosome sizing 155 kb (XC155). The structure of XC155 was determined by restriction analysis and DNA cloning and sequencing of the gel-purified chromosome: it is made of a 'mirror' inverted duplication of the 'right' end of chromosome 1a (approximately 25 kb at each end), and in its central part of a complex tandem amplification of the linearized transfection vector containing the hygromycin resistance gene (over approximately 105 kb). No sequence of the coding region of chromosome 1 (including the 1.6-kb 'switch' region) was found. By contrast, XC155 contains two large (approximately 13 kb) clusters of tandemly repeated subtelomeric sequences (272-bp 'satellite' DNA) as well as telomeric hexamer repeats. This extra chromosome was found to be mitotically stable after >150 generations without selective pressure in vitro. Two sequence elements are considered which may have an effect on mitotic stability and participate to centromeric function in this extra chromosome: the amplification of the input vector and the 272-bp 'satellite' DNA bound by telomeric repeats.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chromosome Deletion
  • Chromosomes / genetics*
  • DNA, Protozoan / genetics*
  • Genes, Protozoan / genetics
  • Leishmania major / genetics*
  • Mitosis / genetics*
  • Recombination, Genetic
  • Telomere / genetics


  • DNA, Protozoan