YAC cloning and identification of rice (Oriza sativa L.) genomic DNA.
Construction of the genomic library by using yeast artificial chromosomes (YAC) has been an approach to a map-based gene cloning strategy. We have obtained more than 2000 YAC clones for the construction of a rice (Oriza sativa L.) genomic library through a procedure that could be summarized as follows: a fractionate by the pulsed-field gel electrophoresis (PFGE), the rice nucleic high molecular weight (HMW) DNA which is partially digested with EcoRI, and recovered fragments larger than 200 kb; ligate the fragments to the EcoRI digested YAC vector pairs pJS97 and pJS98; transform competent spheroplasts prepared from yeast strain YPH252; and select transformants directly on Ura-Trp-double selective media. Southern hybridization results indicated that the sizes of inserts were in the range of 200-820 kb.
Wang
C
C
School of Life Sciences, Zhongshan University, Guangzhou, China.
Wang
J
J
Liu
W
W
Huang
W
W
Liu
L
L
D018244:Chromosomes, Artificial, Yeast; D003001:Cloning, Molecular; D018744:DNA, Plant / Q000302:isolation & purification; D018745:Genome, Plant; D015698:Genomic Library; D012275:Oryza / Q000235:genetics*; D014170:Transformation, Genetic; D015003:Yeasts / Q000235:genetics
D018744:DNA, Plant