Expression and Characterization of a Recombinant Single-Domain Monoclonal Antibody Against MUC1 Mucin in Tobacco Plants

Abstract

A promising alternative to conventional antibodies is the single-domain antibody fragment of the Camelidae (V(HH)), which (because of features such as small length, high expression, solubility, and stability) is preferred to other antibody derivatives. In this report, a recombinant single-domain antibody (V(HH)) against MUC1 mucin in the tobacco plant, which may be considered as a suitable and economical alternative expression system, was produced. This antibody was expressed under the control of a strong constitutive promoter, CaMV35S, and NOS terminator. A plant high-expression sequence (Kozak sequence) was linked at the 5' end for overexpression of the V(HH) gene. The constructed cassette (pBIV(HH)) was transferred to agrobacterium, and the VHH gene was inserted into the plant genome by agrobacterium-mediated transformation. Transgenic lines were selected on kanamycin (100 mg/L) and maintained in soil, and subsequent generations were obtained. The presence and expression of the transgene was confirmed in the transformants by polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and Western blot. Tobacco transgenic lines leave expressed V(HH) at levels varying from 1.12% to 1.63% of the total soluble protein. This report examines the transformation and expression of recombinant single-domain antibody (V(HH)) against antigen-associated tumor in tobacco plants.

Use of Transgenic Oryzacystatin-I-Expressing Plants Enhances Recombinant Protein Production

“…Since no signal peptide was attached to the GR sequence in addition to the CaMV 35S promoter sequence, GR has very likely not migrated out of the cytosol after its mRNA was translated [3]. Recombinant proteins often accumulate only at a very low amount in the cytosol despite that the transgene is efficiently transcribed and the protein is also stable in this cellular compartment [8,10,13,33,37]. Possible reasons are incorrect folding of proteins and the presence of the ubiquitinproteasome proteolytic pathway for recognition and degradation of incorrectly folded proteins [23,46].…”

Comparison of expression systems for the production of human interferon-α2b

“…The prime decision criteria for the choice of an expression system in pharmaceutical production are the safety of the yielded protein in context with the post-translational modification pattern, followed by overall cost, production timescale, scale up capacity, product quality and storage cost [10,12,[27][28][29]. All plant expression platforms, including whole plants systems, have safety benefits over expression platforms based on microbial cells.…”

Nanobody, New Agent for Combating Against Breast Cancer Cells

“…Furthermore, this was the first report of the production of a nanobody against a tumour-associated peptide (Rahbarizadeh et al, 2004a). Nanobodies against this tumour associated antigen (TAA) showed good specificity toward the synthetic peptides and have been successfully expressed in E. coli (Rahbarizadeh et al, 2005), P. pastoris , Tobacco plants (Ismaili et al, 2007;Rajabi-Memari et al, 2006) and CHO cells (Bazl et al, 2007). Moreover, mouse models with breast cancer tumour originated from MCF-7 (human breast cancer cell line) were used for in vivo tumour targeting.…”