Characterization Of A Banana MADS-box Transcription Factor Gene In Transgenic Tomato

Banana is one of the most important fruit in the world, and is ranked as the world’s fourth most important food crop. Being a tropical fruit, banana is a typical climacteric fruit and has poor storage characteristics. Many researches have been focused on banana postharvest, storage physiology and molecular biology.So isolating and analysis of gene function in the fruit of banana’s ripening are both a fundamental biological issue and a highly promising application. With the current development of molecular biology, studies on gene regulation are more in depth, while the transcription factor is an important aspect.An MADS-box gene named MuMADS2 has been isolated from banana fruit by SSH and DNA micro array method of combining and its expression was up-regulated in the early stage of banana fruit after postharvested. Bioinformatics and Microarray analysis show that, MuMADS2 gene encoding the protein as a transcription factor may play a role in the upstream of ethylene signaling. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis shows that MuMADS2 is expressed in the reproductive organs like flowers and fruits,whereas no expression is detected in the vegetative organs like root,stem and leaf. Therefore, the gene may be associated with banana fruit ripening and ethylene biosynthesis.A plant expression vector pCAMBIA1302-MuMADS2 with green fluorescent proteins were transformed into onion epidermal cells by gene gun method. The result of transient expression showed that MuMADS2 gene was located in the nucleus which was in line with the subcellular localization prediction and the characteristics of transcription factors.The plant expression vector pCAMBIA1302-MuMADS2 was transformed into tomato. As a result,37 genetic transformant To plants with hygromycin-resistance have been obtained.The result of PCR test showed that 33 genetic transformant To plants were positive.Selecting plants of obvious phenotypic for Southern blot analysis,the result in the transgenic plants verified that the MuMADS2 has been integrated into the genomes of tomato by both single-copy and multi-copy integration of the target gene.RT-PCR analysis shows that MuMADS2 expresses in roots,stems,leaves,fruits petals and sepals of 53 line of transgenic tomato. The expression lever is higher in roots and leaves than other organs,lower in stems and fruits, no expression is detected in the organs like stem and leaf. GFP detection is also reached the same result.Most of the transgenic tomato have been manifested phenotypes such as showed dwarf, compact growth, internode shortening, leaf folds and pale.Transgenic tomato seeds were significantly lower than the control, and some plants show a high degree of sterility,and soluble solids content was significantly higher than the control. On the base of pCAMBIA1302, MuMADS2 is reverse inserted downstream of 35s promoter, upstream of GFP gene. The constructed vector is named pCAMBIA1302-MuMADS2.