外源赤霉素对太子参块根中细胞分裂素含量及其代谢关键酶基因表达的影响

Effects of exogenous gibberellin on cytokinin content variation in Pseudostellaria heterophylla root and metabolism key enzyme gene expression

【目的】分析外源赤霉素(GA3)对太子参块根中细胞分裂素(CTK)含量变化及其代谢关键酶基因表达的影响,为研究太子参块根膨大发育机制提供理论基础。【方法】将盆栽的太子参幼苗分成4组,其中3个处理组分别根灌150 mg/L GA3(GA3处理组)、20 mg/L多效唑(PBZ)(PBZ处理组)和150 mg/L GA3+20 mg/L PBZ(GA3+PBZ处理组)激素溶液各200 mL,对照组根灌等量清水,在不同块根发育期采集块根样品。运用酶联免疫吸附法(ELISA)检测不同处理时间太子参块根中内源玉米素核苷(ZR)的含量,并采用实时荧光定量PCR(qRT-PCR)分析CTK代谢关键酶基因的表达量变化。【结果】在整个块根发育期,3个处理组太子参块根中的ZR含量总体高于对照组,且呈相同的变化趋势,即从膨大初期到成熟期ZR含量均呈升高—降低—升高的变化趋势,说明外源GA3和PBZ均能促进ZR含量的升高。在GA3处理组中,A-IPT1基因的表达量在膨大初期较对照组高,随着块根的膨大,其表达量呈先升高后降低的变化趋势,直至成熟期低于对照组;tRNA-DMATase基因的表达量在膨大初期较对照组低,随后不断上升高于对照组,呈先升高后降低的变化趋势;A-IPT2基因的表达量在整个太子参块根发育期略高于对照组,而CYP735A、CKX1和CKX2基因表达量均低于对照组。在PBZ处理组中,A-IPT1、tRNA-DMATase和CKX2基因的表达量变化趋势与GA3处理组相反。GA3+PBZ处理组A-IPT2、CYP735A、CKX1和CKX2基因的表达量均明显高于其他组,且与对照组的变化趋势相似。外源GA3可上调A-IPT1和tRNA-DMATase基因的表达水平,下调CYP735A、CKX1和CKX2基因的表达水平,结合ZR含量测定结果可推测CTK在块根膨大初期合成反式玉米素(tZ),而在成熟期合成顺式玉米素(c Z)。【结论】GA3调控CTK的合成和分解是一个动态过程,GA3抑制太子参块根膨大,可能与不同发育时期CTK的代谢有关。

【Objective】Effects of exogenous gibberellin(GA3)on cytokinin(CTK)content variation in Pseudostellaria heterophylla root and metabolism key enzyme gene expression were investigated to provide theoretical basis for studying the root enlargement mechanism of P.heterophylla.【Method】The potted seedlings of P.heterophylla were divided into four groups.Three of the treatment groups were irrigated with 200 mL each of 150 mg/L GA3(GA3 treatment group),20 mg/L paclobutrazol(PBZ)(PBZ treatment group)and 150 mg/L GA3+20 mg/L PBZ(GA3+PBZ treatment group).The control group was irrigated with equal amount of clear water.Root tuber samples were collected at different root development stages.The contents of endogenous zeatin riboside(ZR)in the roots of P.heterophylla during different treatment times were determined by enzyme linked immunosorbent assay(ELASA),the expression changes of CTK metabolism key enzyme gene were determined by real-time fluorescence quantitative PCR(qRT-PCR).【Result】The results showed that the contents of ZR in root tuber of P.heterophylla treated with three treatments was higher than that of control group during the whole root tuber development period,and they all showed the same trend.The change of ZR content showed a rising-falling-rising trend from the early stage of enlargement to the mature stage,which indicated that both exogenous GA3 and PBZ could promote the increase of ZR content.The expression of A-IPT1 gene was up-regulated at the beginning of the enlargement by GA3 compared with control group.With the enlargement of roots,the expression rose first and then decreased,and then it was lower than that of the control group at mature period.The expression level of tRNA-DMATase gene was down-regulated at the beginning of enlargement compared with control and then continuously increased and was higher than the control group,which showed a trend of increasing first and then decreasing.The expression level of AIPT2 gene was higher than control during the whole enlargement period of roots,while the expression levels of CYP735A,CKX1 and CKX2 were down-regulated compared with control group during the whole enlargement period of roots.In PBZ treatment group,the expression variation of A-IPT1,tRNA-DMATase and CKX2 genes were the opposite of those in GA3 treatment group.In GA3+PBZ treatment group,the expression of all genes were greatly higher than other groups,and the variation was consistent with that of the control group.Exogenous GA3 could up-regulate the expression levels of A-IPT1 and tRNA-DMATase genes,and down-regulate the expression levels of CYP735A,CKX1 and CKX2 genes.Combining the detection of ZR content,it was inferred that CTK synthesized trans-zeatin(tZ)at the early stage of root enlargement,and synthesized cis-zeatin(cZ)at the mature stage.【Conclusion】The regulation effect of GA3 on CTK synthesis and decomposition is a dynamic process.GA3 can inhibit the enlargement of P.heterophylla root,which may be related to the metabolism of CTK at different developmental stages.