Plant-based production of recombinant proteins has emerged as an efficient and cost-effective alternative to microbial fermentation and mammalian cell culture systems. Chloroplasts harbor high plasmid copy numbers and can be stably transformed, making them efficient platforms for protein production. In the present study, we used green fluorescent protein (GFP) as a reporter to compare the three major chloroplast promoters (rrn, psbA, and rbcL) involved in protein production in Nicotiana tabacum cv. “Petit Havana.” Three chloroplast transformation vectors were constructed, each regulated by a different promoter, and the transformation was performed via biolistic particle bombardment. Transformants were selected based on spectinomycin resistance and were confirmed by PCR. Among the three promoters, psbA showed the highest transformation efficiency and protein expression levels. Reverse transcription quantitative PCR showed that the mRNA levels (relative to Actin) for psbA (218.21±19.64) were nearly twice that of rbcL (126.60±8.78), and five times that of rrn (43.27±1.57). This transcriptional hierarchy was also observed at protein level. Immunoblotting showed the GFP levels (relative to psbA) were: psbA (1.00), rbcL (0.87), and rrn (0.77), whereas quantification through ELISA revealed relative GFP concentrations of: 616.2±28.7 ng/g LFW for psbA, 510.3±32.4 ng/g LFW for rbcL, and 338.9±100.2 ng/g ng/g LFW for rrn. These quantitative results demonstrate the importance of promoter selection for efficient expression of recombinant proteins in chloroplasts and show that the psbA promoter is suitable for high-efficiency chloroplast expression systems, providing a foundation for advancing plant-based molecular farming.

Wheat (Triticum aestivum L.) is one of the world's three staple crops and accounts for approximately 20% of the total calories consumed by the world's population. It is known that wheat is a difficult crop to introduce foreign genes into, having a large genome (16 Gb) containing three highly related subgenomes (AABBDD). Owing to the low transformation efficiency of wheat, it is difficult to apply new technologies such as genome editing and basic research based on molecular biology, such as the discovery of useful genes and functional analysis. Recently, the completion of a wheat genome map by the International Wheat Genome Sequencing Consortium (IWGSC) and the development of a stable and reproducible wheat transformation system have accelerated research regarding the expression and control of useful genes. In this review, we introduce in detail the recently developed highly efficient Agrobacterium-mediated wheat transformation system and its applications in plant biotechnology, such as RNA interference (RNAi), overexpression, and gene editing using this system.

In recent years, new plant breeding technologies (NPBT) have had enormous effects on breeding and the agricultural industry. In particular, genome editing technology, including site-directed nuclease technologies, has progressed dramatically since the first-generation Zinc finger nucleases to the third-generation clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9). CRISPR/Cas9 technology has yielded a revolutionary breakthrough in the accurate, efficient, and user-friendly genome editing of eukaryotes. Several methods for basic research and applications, such as knock-out, base editing, gene targeting, and transcriptional activation or repression have been derived from CRISPR/Cas9 technology. Herein, we will describe the current progress in NPBTs and also summarize the crops developed by NPBTs. After analyzing the current status of NPBTs and crop development, we have proposed potential strategies for crop development using NPBTs.

Seed dormancy is an important adaptive mechanism to protect seeds under the unfavorable environments. Unlike to wild type species, the seed dormancy trait of cultivated crops has been weakened by breeding programs during the domestication period. Weak seed dormancy often causes preharvest sprouting (PHS) problem in many cereal crops that result in significant economic loss. The seed dormancy is a quantitative trait loci (QTL) controlled by multiple genetic and environmental factors. So far, many QTLs for seed dormancy have been identified from rice and wheat as well as in the model plant Arabidopsis. Unveiling of QTL genes and complex mechanisms underlying seed dormancy is accelerated by the rapid progress of crop genomics. In the present study, we reviewed current status of research progress on the seed dormancy QTLs and correlated genes in Arabidopsis and cereal crops.

Abscisic acid (ABA) is a stress hormone that functions in abiotic stress adaptation in plants. Thus many efforts have been made to identify the molecular mechanisms of ABA signal transduction pathways. Recently there were big advances in understanding molecular mechanisms of ABA dependent expression. From the ABA receptors to the transcription factors, signaling components were discovered and the biological networks among the components were identified. In this review, we describe the ABA signaling components and the rice orthologues identified. These show that signaling network systems of ABA are highly conserved in dicot and monocot plants and we are able to manipulate the ABA signaling components to develop the abiotic stress tolerant crops.