Construction and characterization of a BAC library for flue-cured tobacco line with high resistance to blank shank

Abstract

<p indent="0mm">Tobacco (<italic>N. tabacum</italic>) is an important model crop in molecular biology research. In this study, a bacterial artificial chromosome (BAC) library of a flue-cured tobacco line 14–60 with high blank shank resistance and good quality was constructed. High molecular weight DNA was isolated using intact nuclei from tobacco, partially cleaved with <italic>Hin</italic>d III and cloned into the pIndigoBAC536-S vector. The BAC library consisted of 414,720 clones arrayed in one thousand and eighty 384-microtite plates, with an average insert size of 123 kb ranging from 97.0–145.5 kb. No empty insert clone was found. Based on an estimated genome size of 4500 Mb for common tobacco, the BAC library was estimated to cover 11 times of genome equivalents. The utility of the library was further confirmed by screening the library with the primers of tobacco <italic>hem</italic>A, <italic>NtFT</italic>, and<italic> eIF4E-</italic>1 genes. The high capacity library will serve as a giant genomic resource for map-based cloning of quantitative trait loci or genes associated with important agronomic and smoking quality traits or resistance to blank shank, physical mapping and comparative genome analysis.