Cloning And Expression In E.coli Of Polygalacturonase-Inhibiting Protein Gene On Malus Sieversii F. Neidzweizkyana

Polygalacturonase inhibiting proteins (PGIPs) are extracellular glycoproteins located inplant eell wall. It is believed to play an important role in the defense against plant pathogenicfungi. PGIPs can specially bind and inhibit or reduce the hydrolytic activity ofPolygalaeturonases (PGs)secreted by plant pathogenic fungi,and limit the growth of plantpathogens, as well as eliciting defense responses in plant. Furthermore, PGIPs belong to thesuper family of Leueine-rich repeat (LRR) proteins which also include the products ofseveral plant resistance genes. They are considered to be important candidate genes forgenetic engineering to obtain transgenic plants with increased tolerance to fungal infectionwhich decrease the use of insecticide. Therefore, the basic research of transgenic engineeringfor that purpose is to isolate and identify the PGIP genes.In order to protect and utilize scientificly, it was studied by methods of cloning andexpression of the MsPgip gene of Malus sieversii f. neidzwetzkyana. Sequence analysisshowed that the fragment contains a full coding region of993bp encoding330amino acidresidues with a molecular mass of36.6kD. Its GenBank accession number is JQ001783.Thisdeduced protein has a pI of7.05, a hydrophobic region of24amino acid residues in theN-terminal which was considered to be a signal peptide, two LRRs, and six potentialN-glycosylation sites. MsPgip exhibits a homology of99%‘Granny Smith’、‘Fuji’and‘GoldenDelicious’ apple in amino acid level. The recombinant prokaryotic expression vectorpET-MsPgip was constructed by inserting the cDNA fragment into the prokaryotic expressionvector pET-30a（+）, and then transformed it into E.coli BL21(DE3). The SDS-PAGEdisplays that the expressed protein consistents with the size of expected protein.