Cloning, Transformation and Expression of Human Interferon α2b Gene in Tobacco Plant (Nicotiana tabacum cv. xanthi)

Abstract

BackgroundMolecular farming is the production of important recombinant proteins in transgenic organisms on an agricultural scale. Interferons are proteins with antiviral and antitumor activities and can be used for viral infections and cancers treatments.ObjectivesThis study reports the transformation of INF α2b gene in tobacco plant for the first time in Iran.Materials and MethodsInterferon α2b gene was amplified by PCR using specific primers containing appropriate restriction enzymes, plant highly expression sequence and Histidine tag sequence. Target sequence was cloned in plant expression vector pCAMBIA1304 and the construct named pCAMINFα. pCAMINFα was transferred to E. coli strain DH5α and plated on LB agar medium containing kanamycin 50 mgl-1. The colonies were confirmed by colony PCR and sequencing. The construct was transferred into Agrobacterium tumefaciens by freeze-thaw method and transformed colonies were confirmed by colony PCR. Tobacco plants (cultivar xanthi) were inoculated with A. tumefaciens strain LBA4404 by leaf disc method. Inoculated explants were cultured on MSII (MS + BAP 1mgl-1 + NAA 0.1 mgl-1) at 28°C and darkness for 48 hours. Then explants were transferred to selection medium containing cephotaxime (250 mgl-1) and hygromycin (15 mgl-1) in a 16/8 (day/night) h photoperiod in growth room with an irradiance of 5000 lux. Transgenic plants were regenerated and transferred to perlite. Genomic DNA was extracted from regenerated plants by Dellaporta method at 5-leaf step and transgenic lines were confirmed by PCR with specific primers. Expression of Interferon α2b gene was confirmed by dot blotting.ConclusionsSince no report of interferon alpha production in plants in Iran has been expressed yet, this research could create a field of producing this drug in tobacco, in Iran.

Cloning, Transformation and Expression of Proinsulin Gene in Tomato (Lycopersicum esculentum Mill.)

“…So the use of plant expression systems for large-scale biopharmaceutical production is becoming more accepted (12). Stable transformation is a better choice compared with transient expression since it generates transgenic lines which have the potential to produce millions of seeds (10). In this case plant expression vector pCAMBIA1304 was used which includes the CaMV35s promoter and NOS terminator transcription elements.…”

“…In 1986, transgenic tobacco and calluses of sunflower could produce growth hormone, which can be used in human treatment (8, 9). During the last decades, many different plant platforms have been used to produce recombinant proteins (10-13). Plants based medicines are considered as safe expression platforms for delivery of recombinant medicines for therapeutic use and easy retention and distribution.…”

Multidimensional futuristic approaches to address the pandemics beyond COVID-19

“…Moreover, tests are available to determine if recombinant TPs expressed in plants are bioactive or not. For example, different human interferons having antiviral properties were expressed in plants and shown to be bioactive using human cell lines [ [239] , [240] , [241] , [242] ].…”