Plant & Animal Genome IX Conference

Poster: Large Insert Libraries, Gene Isolation, Etc.
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IDENTIFICATION OF COTTON DISEASE RESISTANCE GENES AND ROOT-KNOT NEMETODE (RKN) INDUCED ROOT-SPECIFIC GENES AS POTENTIAL MARKERS

¹ Dept. of Biochemistry and Molecular Biology, Box 9650, Mississippi State, MS 39762, USA

² Crop Science Research Lab., USDA-ARS, Mississippi State, MS 39762, USA

³ Dept. of Plant and Soil Sciences, Box 9555, Mississippi State, MS 39762, USA

Two approaches were used to identify cotton disease resistance (R) genes and root-knot nematode induced root-specific genes as potential markers from cotton genome. First, a PCR-based strategy was used to isolate cotton R genes. Many plant R genes encode proteins which contain a conserved NBS motif. Two sets of degenerate oligonucleotides were synthesized on the basis of the NBS motif and used as primers in PCR amplification of R genes in cotton. Six different combinations of the two sets of primers had resulted in totally amplifying six different R gene homologs. The nucleotide and derived amino acid sequence data indicate that the six cotton R genes are highly homologous to other plant R genes, including the tomato vascular wilt R gene I2 against soil-borne fungus Fusarium oxysporum, the tomato root knot nematode R gene Mi-1.2 against Meloidogyne incognita, and the Arabidopsis RPS2 gene against Pseudomonas syringae. Cotton I2 gene homologs Gh2-3-1 and Gh3-3-2 were used to identify RFLP polymorphisms between the Near-Isogenic Lines (NILs) differing in Fusarium and Root-Knot Nematode (RKN) resistances. Gh2-3-1 and Gh3-3-2 could be the genes conferring or linked to Fusarium resistance in cotton. Second, the primers, designed from eight amino acid residues of a 14 KDa protein which is specifically only induced in the RKN-infected gall area of the resistant roots after RKN inoculation, were used to clone three cDNA sequences through rapid amplification of DNA ends (RACE). The three cDNAs were named as MIC-1, 2, and 3. Northern analysis indicated that the expression pattern of MIC-3 was similar to that of the 14 KDa protein. MIC1, 2, and 3 cDNA sequences are very similar to each other and all encode a protein of about 14 KDa. Southern analysis revealed that MIC3 was a member of the multiple gene family. An extra RFLP polymorphism band was detected in the RKN resistant NIL. This polymorphism band could be a potential marker linked to the RKN resistance in cotton.