关键词: 苹果属观赏海棠, 叶片, 二氢黄酮醇-4-还原酶基因, 花色苷, 实时荧光定量PCR

Abstract:

Dihydroflavonol-4-reductase (DFR)is the first key enzyme in the later procedure of anthocyanins biosynthesis. With the total RNA drived from leaves of Malus crabapple‘Royalty’ as the template, the full cDNA of DFR gene (1 247 bp) was cloned by reverse transcription polymerase chain reaction (RT-PCR) and rapid-amplification of cDNA ends(RACE).The gene is named as McDFR, containing three open reading frames (ORF, 1 021 bp), and encoding a protein of 348 amino acids. DNA sequence has 2 396 bp, containing five introns, and all the cleave sites obey with the GT-AG rule. The expression of McDFR and the content of anthocyanin and flavonoids were determined respectively by real-time quantitative PCR and spectrophotometer in the mature and young leaves of M.‘Flame’(green young and mature leaf), M.‘Radiant’( red young leaf and green mature leaf), M. ‘Prairifire’ (red young leaf and green mature leaf)and M.‘Royalty’( purple young and mature leaf). The results showed that McDFR was expressed in both mature and young leaves of the above four cultivars.The trends of anthocyanin content were consistent with that of the relative expression of McDFR gene. The expression levels of McDFR in the young leaves were higher than that in the mature leaves. Among the four cultivars, the M. ‘Prairifire’ in the young leaves showed the highest expression levels of McDFR. Flavonoid content in young leaves was higher than that in the mature leaves, and the expression of McDFR was also higher than in the mature leaves. In the mature leaves, except for the green leaf variety M.‘Flame’, the higher content the flavonoid the higher expression level McDFR .These results have determined that McDFR plays an important role in anthocyanin and flavonoid metabolic pathways and the color formation in the leaves.

Key words: Malus crabapples, leaf, McDFR, anthocyanin, real-time quantitative PCR