Eucalypts are a major forest resource globally and the area of eucalypt plantations for pulp and paper production is expanding rapidly in Australia. Consequently, there is an increasing need to breed eucalypts with improved wood properties. Since many high value wood traits are under strong genetic control, identification of DNA markers linked to these traits will have application in breeding programs. In recent years there has been a shift in marker strategy away from QTL mapping in pedigrees to association studies in unrelated populations. In the latter approach, single nucleotide polymorphisms (SNPs) in candidate genes are screened to identify SNPs that significantly associate with wood traits. Significant SNPs could be used for marker-assisted selection (MAS) in breeding programs. The objectives of this study were to identify candidate genes that may influence pulp yield in eucalypts and to identify SNP variants in those genes that associate with superior wood and pulp traits. / Approximately 300 trees from a full-sib Eucalyptus nitens progeny derived from a wide intra specific cross were used for gene discovery. DNA microarrays containing ~5800 young xylem of cDNAs Eucalyptus grandis were screened with probes synthesised from RNA isolated from trees with either high or low pulp yield. Forty-six transcripts were differentially regulated, of which 27 were more abundant in high pulp trees and 19 were more abundant in low pulp trees. All differentially expressed cDNAs were partially sequenced and searched against existing gene databases. Six genes were selected as putative pulp yield candidate genes based on their significant similarity to genes with known function and were named EgrCesA3 (cellulose synthase), EgrNAM1 (NAM family protein), EgrXET (xyloglucan endotransglycosylase), EgrGalk (galactokinase), EgrHB1 (class III homeodomain leucine zipper protein) and EgrZnf1 (C3HC4 type zinc finger protein). / Real-Time PCR was carried out on selected genes to confirm the accuracy of the microarray results. Full length cDNAs were obtained for EgrCesA3, EgrHB1 and EgrZnf1 and the candidate genes were partially characterised. An additional candidate gene, the novel gene EgrPAAPA, was selected based on previous research due to its high expression in the cambium and its expression in eucalypt branches. EgrPAAPA was cloned by screening an E. grandis cDNA library and fully sequenced. The full length EgrPAAPA encodes a short 172 amino acid protein rich in alanine, glutamic acid and proline residues. The EgrPAAPA protein appears to be a hydroxyproline-rich glycoprotein (HRGP) and the repetitive ‘PAAPA’ motif suggests that it might play a structural role in cell wall development. Southern blot analysis revealed that E. grandis has a single copy of the EgrPAAPA gene and northern blot analysis revealed that EgrPAAPA is most strongly expressed in xylem tissues. / Allelic variation in EnCesA3, EnNAM1, EnPAAPA and EnHB1 was examined by sequencing each gene in 16 to 24 unrelated E. nitens individuals. SNPs were identified by sequence analysis and patterns of nucleotide diversity, linkage disequilibrium and the selection of suitable polymorphisms were estimated. A moderate level of nucleotide diversity (θw = 0.0056 and π = 0.0039) was observed and linkage disequilibrium was generally low, extending only a few hundred base pairs in each gene. Negative selection has been operating in EnHB1. Selected TagSNPs from EnNAM1, EnHB1 and EnPAAPA were genotyped across 300 unrelated E. nitens trees which had been phenotyped for six wood quality traits including pulp yield, cellulose, lignin, Klason lignin, microfibril angle (MFA) and density. Five highly significant genetic associations (p<0.01) were detected between several SNPs in EnHB1 and all wood quality traits except density. A significant association was also found between EnPAAPA and MFA (p<0.05). No significant associations were found with any of the EnNAM1 SNPs. The strong genetic associations between SNPs in EnHB1 and a range of wood traits is consistent with this gene’s known role as a transcription factor controlling vascular development. Validation of these associations in different populations will be necessary in order to confirm these results. Alternatively, QTL mapping can be performed in order to confirm whether QTL for wood property traits can be detected at the EnHB1 and EnPAAPA loci.
| Identifer | oai:union.ndltd.org:ADTP/245136 |
| Date | January 2008 |
| Creators | Bhuiyan, N. |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
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