Research Interests
Application of functional genes for postharvest improvements on fruit and vegetable crops via transgenic technology and biosafety evaluation via genomic and proteomic profile assays in transgenic plants.

1. I.Transgenic biotechnology toward post-harvest improvements for broccoli.
• In our laboratory, crop biotechnology including the development of molecular markers and genetic transformation for crop improvement were our previous foci in the past few years. Since postharvest biotechnology is believed to be an essential issue for the reduction of harvested loss and maintenance of higher product quality in the vegetables, fruits and ornamental crops. With the rapid growing in knowledge
  of functional genomic studies, we now have a great pool of genes for choice for molecular breeding. It is obvious to say that transgene technology will play an indispensable role in crop improvement in the future. From our previous experiences, we have development a stable transformation system in broccoli. We are now using this system as a model for the evaluations on genes related to the postharvest senescence.
• Broccoli is a highly perishable fresh vegetable and tends to turn yellowing when it is subjected to ambient temperature in 1 to 3 days. Our previous studies on the transformation of broccoli with anti-senescence genes have succeeded to obtain transgenic lines with cytokinin synthase [ipt (isopentenyl transferase) ] gene, mutant ethylene response sensor gene(boers), anti-chlorophyllase (Anti-CHL-1) gene(Fig.1)

  Fig. 1:Comparison on postharvest yellowing retardation in progeny of anti-chlorophyllase 1(BoAnti-sense CLH-1) transformants and non-transformed controls in detached leaves(a) and floral heads(b) postharvest and stored at 20℃

  
  and anti-fungal protein(AFP-3) gene(Fig.2) respectively. The ipt transformed lines did have the effect on the retarding of floret yellowing in harvested floret branchlets without any sever effect on morphological characters. The Boers and Anti-CHL-1 transgenes also showed the retarding effects on postharvest yellowing, but some drawbacks with the constitutive promoter were observed in most transgenic lines. AFP-3 transgenic broccoli also indicated the possibility of resistant to Rhizoctonia solani. Further transformation on some potential candidate genes such as pHellsgate-RNAi-BoBFN1 (bifunctional nuclease gene)、Cor15a-CBF1(C repeat/dehydration responsive element binding factor 1)、CaMV35S-CBF1 and Cor15a-ipt for their possible effects on postharvest improvements are under evaluation.

Fig. 2: Northern and RT-PCR expression assay on the anti-fungal protein transgenic broccoli.



2. II.Exploring the possible unintended effects of transgenic broccoli via genomic and proteomic Studies.
• Our previous studies on the transformation of broccoli with cytokinin synthase [ipt (isopentenyl transferase) ] gene together with senescence associated promoter(SAG) and NPTII gene had obtained some potent ipt transgenic lines with retarding of floret yellowing in harvested floret branchlets. Among those, three T5 ipt transformation selfed-lines(93-101,102,103) with good agronomic performance and showing the retarding effect on yellowing in harvested floret heads have been selected. According to the “Evaluations of food safety of genetically modified food” announced by Department of Health, Executive Yuan, ROC, we need to process safety evaluations, including the preparation of genetically modified food and the toxicity, allergy, nutrient components, and antibiotics of genetically modified food before the release for commercial use. A collaborative and integrated project entitled “Establishment on the food safety evaluations on the transgenic ipt(isopentenyl transferase) broccoli” has included four major aspects:1) Exploring the possible unintended effects and genetic stability from studies of genomic and proteomic profiles in transgenic ipt(isopentenyl transferase) broccoli, 2) evaluation of the food safety issues of ipt transformed broccoli lines by a animal-feeding model, 3) investigation on specific field characteristics and essential nutrient components in transgenic ipt(isopentenyl transferase) broccoli lines, and 4) assay of key composition, allergenicity and toxicity of ipttransformed broccoli lines. In our laboratory, we have investigated the genetic stability, transgene activities and the possible occurrence of un-intended effects via molecular biology tools through the proteomic profile and genomic DNA microarray studies(Fig. 3).

Fig.3 Exploring the possible unintended effects of transgenic broccoli via (A) DNA microarray and (B) 2-D protein profile comparisons.

Selected Publications

Full publication list

• Chen, L. F. O., Hwang, J. Y., Charng,Y. Y., Sun , C. W., and Yang, S. F. (2001). Transformation of broccoli (Brassica oleracea var. italica) with isopentenyltransferase gene via Agrobacterium tumefaciens for postharvest yellowing retardation. Mol. Breed. 7: 243-257.
  
  
• Lai, Y. C. and Chen,L. F. O.(2002). Flow cytometric analysis of nuclear cell cycle phases in relation to plant regeneration in Petunia hybrida. J. Genet. Mol. Biol. 13:13-20.
  
  
• Chen, L. F. O., Yang, S. F. and Huang, J.Y. (2003). High Efficiency Plant Transformation of Brassica olercea. Patent No.: US 6,667,428 B1.
  
  
• Lo, H. F., Chiu, S. H., Lo, S. F. and Chen, L. F. O. (2004). Random amplified DNA disclosed Genomic instability in successive cutting-propagated clones of sweet potato(Ipomoea batatas (L.) Lam.) with N-fertilizer treatments. J. Genet. & Mol. Biol. 15:45-57.
  
  
• Chen, L. F. O., Huang, J. Y, Wang,Y. H., Chen, Y. T. and Shaw, J.F. (2004). Ethylene insensitivity and post-harvest yellowing retardation in mutant ethylene response sensor(boers) gene transformed broccoli(Brassica olercea var. italica.) Mol. Breed.14:199-213.
  
  
• Lin, K.H., Tsou, C.C., Huang, S.Y., Chen L.F.O. and Lo, H.F. (2006). Enhanced flooding-stress tolerance in sweet potatoes after pacloburazol treatment. J. Plant Physiol. 163:750-760.
  
  
• Chen, L. F. O., Lin, C. H., Kelkar, S. M., Chang, Y. M. and Shaw, J. F. (2007). Transgenic broccoli (Brassica oleracea var. italica) with anti-sense chlorophyllase (BoCLH1) on delaying postharvest yellowing. Plant Sci. 174:25-31.