Flow sorting and microcloning of maize chromosome 1

Hereditas
Lijia LiYunchun Song

Abstract

Flow sorting maize chromosome 1 and construction of the first chromosome 1 DNA Lambda library are described. Maize metaphase chromosome suspensions were prepared from synchronized seedling root tip cells of the maize hybrid line Seneca 60 and stained with propidium iodide for flow karyotyping and sorting. The observed flow karyotype was very similar to the predicted flow karyotype constructed based on published values for the relative chromosome sizes of Seneca 60. The estimated size of chromosomes from the peak for the chromosome 1 matched the expected size of maize chromosome 1. The peak for the chromosome 1 was well resolved from other peaks on the flow karyotype. An average of 7 x 10(3) chromosomes of chromosome 1 could be produced from 10 root tips. About 0.6 million chromosomes of maize chromosome 1 were sorted and pooled based on the cytogram of fluorescent pulse area Vs fluorescent pulse width and stored at -20 degrees C in the freezer. DNA isolated from sorted chromosomes was good quality of more than 100 kb in size. Chromosome 1 DNA was partially digested with BamHI, dephosphorylated and ligated with arms of BamHI digested Lambda Dash vector. A total of 1.2 x 10(5) independent recombinants with the average insert size...Continue Reading

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Citations

Aug 17, 2012·Functional & Integrative Genomics·Jaroslav DoleželHana Simková
Jan 11, 2014·Biotechnology Advances·Jaroslav DoleželHana Simková
Aug 22, 2012·Human Vaccines & Immunotherapeutics·Justyna BazanAndrzej Gamian
Jul 25, 2018·Current Protocols in Plant Biology·Suchitra ChavanJulia Frugoli

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