Construction and analysis of a subtracted cDNA library of Betula platyphylla female inflorescence

Female inflorescence of Betula platyphylla was sampled at an interval of eachtwo days to analyze the background of gene expression in floral phase. On the basis of SMARTstrategy, the driver cDNA was obtained from total RNA of the last sample and the tester cDNA wasfrom that of the others by RT-PCR which were subsequently used to construct a subtracted cDNAlibrary. The result of the ESTs (expression sequence tags) blastX showed that the genes in thesubtracted cDNA library could be mainly clustered into 5 groups related to metabolism,transportation and signal transduction, cell cycle, stress response, and regulation. Therelationship between gene expression and development was also discussed.

Responses of Wheat (Triticum aestivum) to Grain Aphid (Sitobion avenae) Infestation and Mechanical Wounding Using a cDNA Subtractitve Library Approach

Aphids are major insect pests of cereal crops, acting as virus vectors as well as causing direct damage. The responses of commercial wheat (cv. Claire) to grain aphid (Sitobion avenae) infestation and mechanical wounding were investigated in this study, with the aim to eventually identify a source of molecular markers to breed wheat for enhanced insect resistance, and in particular for enhanced resistance to phloem-feeding insects. Mechanical wounding was included in this study as a comparison with aphid feeding to distinguish between insect-specific responses in wheat plants to those involved in a general wounding response. Wheat (Triticum spp.) is known to have partial resistance toward aphids [1]. The plant response and defence against insect feeding are complicated, but always follow the same principle: insect detection, signal transmission to initiate defence, changes in plant gene expression and subsequent production of defensive compounds, which may be targeted to the wound site to deter or kill insects. Defensive gene products/proteins reach the target area and deter or kill insects. Whether the last step is successful or not depends on the resistance and susceptibility of the plant towards that particular pest. In the light of this principle, it is important to detect changes in gene expression, first at the transcriptional level, which is useful for detection of early-stage responses, and then once sufficient time is allowed for the plant to produce defensive gene products, responses at the proteome level can be identified. Work presented in this study focuses on the changes at the transcriptional level;differential responses at the proteome level were investigated and presented in Ferry et al. 2011 [2] and Guan et al. 2015 [3]. Two cDNA subtractive hybridization libraries were constructed, one to identify transcripts involved in the responses to aphid infestation, and the second to identify transcripts involved in responses to mechanical wounding. Following subtractive hybridization, 520 and 800 clones were obtained from the subtractive hybridization between aphid-infested and un-infested wheat cDNAs and between mechanically wounded and un-wounded wheat cDNAs, respectively. Over 70% of the total clones were sequenced and 44% and 55% of sequenced clones were successfully identified by homology to known sequences held at NCBI with Blastx search engine in aphid-infested vs un-infested and mechanically wounded vs un-wounded cDNA subtractive libraries, respectively. These results reveal that the differences in the response of commercial wheat (cv. Claire) plants towards aphid infestation and mechanical wounding are subtle. Although the majority of differentially expressed putative genes after aphid infestation or mechanical wounding were involved in metabolic processes and photosynthesis, the majority of the genes expressed were different. Genes encoding glutathione transferase (GST), apoptosis and proteolysis were up-regulated after aphid feeding, suggesting their importance towards plant defence/tolerance against aphid attack. These results suggest that commercial wheat does have a certain degree of tolerance to aphids, but appears to lack a specific response to aphids;these findings are supported by those presented in Ferry et al. 2011 [2].

Identifcation and Characterization of 177 Unreported Genes Associated with Liver Regeneration

The mammalian liver has a very strong regeneration capacity after partial hepa- tectomy (PH). To further learn the genes participating in the liver regeneration (LR), 551 cDNAs selected from subtracted cDNA libraries of the regenerating rat liver were screened by microarray, and their expression pro?les were studied by cluster and generalization analyses. Among them, 177 genes were identi?ed unre- ported and up- or down-regulated more than twofold at one or more time points after PH, of which 62 genes were down-regulated to less than 0.5; 99 genes were up-regulated to 2–10 folds, and 16 genes were either up- or down-regulated at dif- ferent time points during LR. By using BLAST and GENSCAN, these genes were located on responsible chromosomes with 131 genes on the long arms of the chro- mosomes. The cluster and generalization analyses showed that the gene expression pro?les are similar in 2 and 4, 12 and 16, 96 and 144 h respectively after PH, suggesting that the actions of the genes expressed in the same pro?les are similar, and those expressed in di?erent pro?les have less similarity. However, the types, characteristics and functions of the 177 genes remain to be further studied.