Plant & Animal Genome VI Conference

P20

HIGHLY EFFICIENT CONSTRUCTION OF A RICE BAC LIBRARY USING VARIETY MINGHUI 63 FOR CLONING GENES OF AGRONOMIC IMPORTANCE

1. National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China
2. Crop Biotechnology Center, Texas A&M University, College Station, TX 77843, USA

"Minghui 63" is the restorer line for a number of the most important hybrids in rice production in China; hybrids developed using this variety occupy a total of over 7 million ha per year in the last decade. Besides its extraordinarily high combining ability in yield, it also possesses many desirable characteristics such as wide adaptability, good grain quality and resistance to a number of races of bacterial blight and blast. Using Minghui 63 as a parent, we have conducted molecular marker-based genetic mapping of several important genes including the ones governing grain quality, fertility restoration, photoperiod sensitive male sterility and bacterial blight resistance. We also investigated the genetic basis of heterosis using hybrid "Shanyou 63" having Minghui 63 as one of the parents. To facilitate long-term commitment in genetic analysis and molecular cloning of the superior genes in the genome of Minghui 63, we have constructed a large fragment library using bacterial artificial chromosome (pBeloBAC 11) as the vector. Size fractionated Hind III digest of genomic DNA was ligated to the BAC vector, and the ligation mixture was used to transform the bacterial strain DH10B. A total of over 20,000 clones were obtained with the average insert size of ca. 150 kb, ranging from 90 to 240 kb. The clones thus obtained represent a seven-genome equivalent. There is a major improvement in the techniques of library construction which resulted in 20-fold increase in the transformation efficiency. The library is now being used for physical mapping of several genomic regions for map-based cloning.