Agrobacterium tumefaciens-MEDIATED IN-PLANTA TRANSFORMATION OF INDONESIAN MAIZE USING pIG121Hm-Cs PLASMID CONTAINING nptII AND hpt GENES

Abstract

Maize ( Zea mays L.) productivity in Indonesia is challenged to be increased using genetic engineering. Recent advances in Agrobacterium tumefaciens -mediated in-planta transforma-tion makes it possible to transform maize with low cost, and simple method. This study aimed to confirm pIG121Hm- Cs plasmid in A. tumefaciens , and to estimate the efficiency level of A . tumefaciens -mediated in-planta transformation of Indonesian maize by using pIG121Hm- Cs plasmid containing npt II and hpt genes. A series of studies were conducted including confirmation of gene construct of pIG121Hm- Cs plasmid in A. tumefaciens , transformation of four maize lines through A. tumefaciens -mediated in-planta technique, acclimatization of transformant plants and molecular analysis of selected plants using polymerase chain reaction (PCR). The pIG121Hm- Cs plasmid was confirmed via PCR analysis using specific primers of npt II and hpt genes and resulted 700 bp and 500 bp for fragments of npt II and hpt , respectively. After selection, acclimatization and molecular analysis steps, the efficiency levels of transformation of four maize lines were low, ranging from 3.8% to 12.8%. The level of transformation efficiency of ST-27 line was the highest accounting for 12.8% of 45 planted embryos on selection medium based on PCR analysis using specific primer for npt II gene. Overall, A. tumefaciens -mediated in planta transformation on maize floral pistil in this study proved to be successful and rapid. Therefore, this enhanced transformation method will be beneficial for Indonesian maize genetic engineering.