Antibiotic-Free Chloroplast Genetic Engineering

Since chloroplast genetic engineering has the advantages of high gene expression and no gene contamination, it is emerging as a new technology to replace plant genetic engineering, which overcomes many concerns of nuclear genetic engineering. However, transformation of transgenic higher plant chloroplasts requires the introduction of thousands of copies of the transgene into the chloroplast genome with stable integration of the antibiotic resistance gene as a selectable marker, which will amplify the antibiotic resistance gene. The current concern is that antibiotic resistance genes may be transferred to weeds or disease-causing microorganisms in the gastrointestinal tract or soil, making them resistant to treatment with such antibiotics. Additionally, there is public concern that plants containing antibiotic resistance genes may inactivate oral doses of antibiotics. Therefore, there is a need to develop effective antibiotic-free chloroplast transformation strategies to eliminate the possibility of their transfer to microorganisms or plants and to alleviate public concerns about GM crops.

Fig. 1. Deletion of antibiotic resistance or marker genes from chloroplast transgenic plants. (Daniell H, et al., 2001)

Services

Over the years, the team at Lifeasible has accumulated rich experience in chloroplast genetic engineering, dedicated to developing multiple effective strategies to provide solutions for antibiotic resistance genes. Our aim is that designing transgenic crops based on a chloroplast gene transformation platform without the use of antibiotic resistance genes should eliminate their potential risk of transfer to the environment or gut microbes. According to your project needs, we will choose the best strategy for chloroplast transformation without antibiotics.

• Selection Without Using Antibiotic Resistance Genes in Chloroplast Transformation

We performed selection without antibiotic resistance genes using the spinach betaine aldehyde dehydrogenase (BADH) gene as a selectable marker to obtain rapid regeneration of chloroplast transgenic plants.

• Removal of the Antibiotic Resistance Genes in Chloroplast Transformation

After generation of chloroplast transgenic plants, we employ advanced biotechnology to achieve efficient removal of antibiotic selection markers.

Methods for Antibiotic-Free Chloroplast Genetic Engineering

We employ homologous recombination and segregation pathways that act on the chloroplast genome and are based on direct duplication, transient co-integration or co-transformation, and segregation of trait and marker genes. In addition, we employed exogenous site-specific recombinases and their target sites to remove marker genes from plastids. Currently, our engineers have successfully used a variety of methods to eliminate antibiotic resistance genes in nuclear transgenic crops:

• Co-transformation.
• Loop-out recombination.
• CRE/Lox system.
• BADH and selection of genes using transient co-integration.

Lifeasible can meet the needs of customers on time and on budget through a wide range of chloroplast genetic engineering strategies. Our aim is to be customer-centric and to provide the highest quality service to customers around the world. Our skilled and dedicated scientific researchers ensure that the most appropriate methods and techniques are selected for each specialized chloroplast project. Our customer service representatives are enthusiastic and trustworthy 24 hours a day, 7 days a week. If you are interested in our services, please feel free to contact us for more information or a detailed discussion.

Reference

1. Daniell H, Wiebe P O, Fernandez-San Millan A. (2001) Antibiotic-free chloroplast genetic engineering–an environmentally friendly approach[J]. Trends in Plant science. 6(6): 237-239.