In Silico Cloning and Transcript Level Analysis of PpPFK in Peach Fruit Stored at Different Low Temperatures
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Abstract:
In order to clone PpPFK(Prunus persica phosphofructokinase) gene and analyze its transcript levels of peach fruit stored at different low temperatures, PpPFK gene complete cDNA length was isolated with in silico cloning method and RT-PCR (reverse transcription PCR), and its transcript levels of peach fruit stored at 0 ℃ and 5 ℃ were examined through reverse transcription real-time quantitative PCR (RT-qPCR). The PpPFK gene with 1405 bp was gained, which codes 232 amino acids with an ORF (open reading frame). The ORF check test was conducted and the result suggested that the sequence of PpPFK gene through PCR was the same as that in silico cloning. The amino acid sequence of PpPFK gene in peach fruit was very similar to that of Glycine max and Vitis vinifera. The RT-qPCR analysis of PpPFK gene indicated that its transcript levels of peach fruit stored at 5 ℃were higher than that at 0 ℃ during first 14 days, resulting in an obvious chilling injury of peach fruit stored at 5 ℃. These results revealed that PpPFK was cold response gene, and differential responses existed in peach fruit stored at low temperatures, indicating its important role in low temperature stress response.
