Dissection and high-yield recovery of nanometre-scale chromosome fragments using anatomic-force microscope

Abstract

We developed a new method to dissect and recover nanometre-scale chromosome fragmentsfrom specific regions of a chromosome by atomic-force microscopy (AFM) witha novel AFM cantilever control. Recovery of the chromosome fragments usingthis method was highly reproducible, with a success rate greater than 90%. Thewidth of the dissection depended on the cantilever-tip diameter; similar widths(240 ± 92 nm on average) were obtained when a standard silicon cantilever was used. We alsoachieved successive dissection and recovery of five fragments located on a specific regionof a chromosome. Rinsing the specimen with a biological buffer, such as PBS,facilitated the recovery. This treatment may cause the chromosome structures to bemore relaxed. The method, called the ‘scratch method’, will be a useful tool toeasily obtain nucleotide sequences with positional information on a specific regionof a chromosome and will contribute to various fields of genetic investigations.