Microdissection and painting of asparagus L5 chromosome

Asparagus officinalis L. is a dioecious plant. A region called the M-locus located on a pair of homomorphic L5 chromosomes controls sexual dimorphism in asparagus. This study aims to conduct microdissection and DOP-PCR on the L5 chromosome, and then physically locate the DOP-PCR products of the L5 single chromosome on the metaphase chromosome of asparagus by chromosome painting. The micromanipulator results indicated that the L5 chromosome was successfully microdissected and its DNA was amplified by DOP-PCR. The molecular weight of its PCR products ranged from 250 to 3000 bp according to 1% agarose gel electrophoresis. The second round DOP-PCR products of the L5 chromosome were labeled with Alexa Fluor-488 by using the nick translation method and the metaphase chromosomes of male asparagus were painted, showing that the fluorescence signals were distributed on the centromeres and secondary constriction areas of all chromosomes. This study has provided a basis for the construction of L5 chromosome DNA libr...