title: [Construction of cDNA library of Magnaporthe grisea with magnetic bead].
abstract: OBJECTIVE
  We constructed cDNA library of Magnaporthe grisea. The good quality cDNA library could facilitate finding proteinaceous elicitors of M. grisea, and elucidating the mechanisms of the M. grisea--rice interaction.
  
  
  METHODS
  The Oligo(dT) combined with the magnetic bead was used to extract mRNA from total RNA of Magnaporthe grisea and as primers to synthesize the first-strand cDNA. Terminal transferase introduced PolyA into 3'terminal of the first cDNA strand, then the PolyA was used for amplifying the second-strand cDNA. Restriction enzyme and adapter were avoided in this research, which could solve technical limitation of the traditional method. Because all reactions were done in one centrifuge tube, this process could reduce the risk of cDNA loss and cross-contamination. The primers designed in this research could clone the amplified cDNAs into expression vector in a desirable orientation.
  
  
  RESULTS
  The cDNA library constructed had a high titer of 8.9 x 10(6) cfu/mL, and contained a total clones of 8.9 x 10(7) cfu, with an average inserts size of about 1380 bp.
  
  
  CONCLUSIONS
  Constructing cDNA library with magnetic bead was a highly efficient method using only small amount of experimental materials within a short period.
authors: College of Life Science, Yangtze University, Jingzhou 434025, China. ziwu211@126.com
authors: Xu
authors: X
authors: Feng
authors: Wu
authors: W
authors: Xiaoli
authors: Qiu
authors: Q
authors: Dewen
mesh_terms: D018076:DNA, Complementary / Q000235:genetics; D015723:Gene Library; D020082:Magnaporthe / Q000235:genetics*; D008280:Magnetics; D008863:Microspheres; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D016133:Polymerase Chain Reaction; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification
chemical_list: D018076:DNA, Complementary; D009838:Oligodeoxyribonucleotides; D012331:RNA, Fungal; D012333:RNA, Messenger; C027903:oligo (dT)