Utility and construction of full-length cDNA library of Malus hupehensis post-introduced with salicylic acid.

In this experiment,total RNA was extracted from leaves of Malus hupehensis treated with SA by improved CTAB method and mRNA was purified.The full-length cDNA library was constructed using the SMARTTM PCR cDNA Synthesis Kit,from which PGIP gene was cloned.The results showed that the total RNA was non-degradable,non-polluting.mRNA dispersion were mainly concentrated in the 500～2 000 bp,and there was no rRNA residues.Dispersion of ds cDNA was mainly distributed between the 300 and 2 000 bp.PCR fragment size was between 200 and 2 000 bp,indicating that the quality of ds cDNA synthesized was better and we successfully constructed a full-length cDNA library.PGIP resistance gene was cloned by PCR from this cDNA library,named as MhPGIP,and GenBank accession number was FJ449708.The homology of nucleotide sequence and amino acid sequence with the apple was 98% and 97% respectively.Two leucine-rich repeat sequences were found in the MhPGIP sequence.The constructed library provides resources for finding,cloning and utilizing new disease-resistant gene,and basis for studying the mechanism of resistance of apple in the future.