Characterization of transgenic tobacco plants carrying small subunit ADPglucose pyrophosphorylase cDNAs from sweet potato

Abstract

In order to test a possibility of forming an active ADPglucose pyrophosphorylase, (AGPase) by homotetrameric composition of single subunit in plant, transgenic tobacco plants were generated with two previously isolated cDNAs (sTL1 and sTL2) encoding the small subunit of sweet potato AGPase. Transformation was carried out with a mixture of the binary vectors, pMBP 1-sTL1 and -sTL2, under the control of CaMV 35S promoter. The PCR analysis of the randomly selected fifteen transgenic plants showed that ten transgenic lines were transformed by sTL1 and seven lines by sTL2. Only two transgenic lines were transformed with sTL1 and sTL2. The integrated sTL1 and/or sTL2 were actively transcribed. The steady-state transcript levels of the transgenic plants were higher than those of nontransgenic plants. The sTL1-transformed lines showed higher transcript levels than sTL2-transformed lines did although the transcript levels were variable among transgenic lines. Despite the differential expression of AGPase mRNA between nontransgenic and transgenic plants, there was no significant difference in the activities of AGPase, indicating no sweet potato AGPase activity in transgenic tobacco plants. The immunoblot analysis of the small subunit polypeptide showed that the protein levels remained unchanged in transgenic plants. The overexpressed transcripts of sweet potato AGPase small subunit in tobacco plants do not seem to produce an active form of AGPase.