Gene Cloning And Expression Of Rubisco Activase In Wheat

Although Rubisco possesses half of the soluble protein in plant leaf, its carboxylation efficiency is still low, because only the active form of Rubisco is capable of catalyzing CO2 fixation .The sugar phosphates such as RuBP can tightly bind to the active site of Rubisco, causes conformational changes of the enzyme, and produce an inactivated dead-end complex. Rubisco activase (RCA) is a soluble chloroplast protein that is coded in nucleus and has the character of ATPase. RCA urges RuBP dissociate from the active site of Rubisco, switches Rubisco from an inactive to an active form by the ATP-dependent release of tight-binding sugar phosphates and so keeps up the initial activity and total activity of Rubisco. A cDNA fragment encoding RCA was cloned from a cDNA library constructed with mRNA from wheat leaves, and the function of RCA during dark-induced aging was investigated. The main results were as follows:Special primers of T7 and T3 was designed according to pBK-CMV phagemid vector, and objective gene was cloned through PCR. The rca gene fragments were cloned and registered in GenBank (AF206372);According to the information about ??subunit of RCA in other plants obtained from GenBank, one important conserved amino acid sequences deduced in the ?? subunit of wheat RCA was confirmed. With midst rca fragment cloned from cDNA of wheat leaves as a probe template, northern analysis was done in dark-induced senescent wheat leaves and control. The expression pattern of rca mRNA was observed. The Pn,CE,Fv/Fm,the total activity of Rubisco and chlorophyll content were measured in dark-induced senescent and control wheat leaves. The factors that were decisive in the decline of Pn and the function of RCA during aging were discussed.