Fruit Ripening-associated Gene Expression In Carya Cathayensis

Carya cathayensis is an important oily woody species and the oil content of its fruit is more than 70%. The oil content of this plant exceeds that of "Caoyou No.2", a rape line (B. campestris L.) and oil palm, king of oily plants, while most of the oil is unsaturated fatty acids. Thus, it is inferred that there may be some special mechanism for oil synthesis. With 13 samples collected from early August to September when it is the period of fruit ripening, an important stage for accumulation and conversion of stored substances of the fruit, ripening-related gene expression has been studied using DDRT-PCR and semi-quantitative RT-PCR. The main results are as follows:1. With an improved CTAB method, integral, quality, highly pure RNA was extracted, which is a choice for RNA extraction from the seeds of other oily woody species.2. 473 differentially expressed fragments were got from DDRT-PCR products, 130 out of which failed in re-amplification.3. The above differentially expressed fragments were cloned and sequenced. With the exception of 10 sequences that are contaminated by vectors, 66 cDNA fragments were found to be involved in cell structure, cell resistance and defense, protein synthesis and processing, energy metabolism and signal transduction, out of which 30 fragments were found to be highly homologous to fatty acid metabolism-related genes viz. caleosin, lipase, fatty acid desaturase, acyl-CoA binding and malate synthase. In addition, 110 cDNA fragments were found to have high identities to known unigenes, and 157 cDNA fragments didn't match any sequence in the nr-database.4. Semi-quantitative RT-PCR was employed to validate 10 differentially expressed fragments obtained from DDRT-PCR (e.g. fatty acid metabolism, signal transduction and energy metabolism), which have a good repeatability in DDRT-PCR and are of significance by alignment to study temporal and spatial expression of relevant genes. As a result, the result obtained from DDRT-PCR was similar to that obtained from Semi-quantitative RT-PCR in gene expression profile except that of malate synthase.5. The final results show that most of fatty acid metabolism-related genes were got from August 11th to 23rd. Thus, it is postulated that this period of time may be the key period of oil accumulation.