Correlation of Cry1Ac mRNA and protein abundance in transgenic Gossypium hirsutum plant

Abstract

Transcription and translation in eukaryotes are distinct processes of the molecular cascade leading to protein production from genetic material. However, establishing correlation between mRNA expression and protein abundance, the end results of the two processes of central dogma, remains a challenge. For transgenic plants, such correlation between mRNA and protein expression serves as a guide to design the transgene, in particular the choices of promoter and codon usage to ensure stable expression of the target protein in relevant tissues under various stress conditions. To elucidate level of mRNA-protein correlation in a commercial transgenic cotton plant Gossypium hirsutum, Bollgard II^® (MON15985), we present the results of Cry1Ac protein expression correlating with corresponding mRNA levels. Protein was quantitated using a home-grown validated ELISA assay with a monoclonal-polyclonal antibody pair, whereas mRNA level was detected by a real-time quantitative PCR assay using standardized reference genes. Our results indicate that protein and mRNA levels are highly correlated in the leaves, but not in squares and stem. The correlations seem to be consistent between young and mature leaves and increase over time of harvesting of samples from months 1-3. These findings demonstrate that transcript level measurement could serve as a proxy to protein abundance for this commercially important cotton species, particularly for leaf tissues which are the most vulnerable organs to cotton bollworms and other pathogens.

Supplementary information: The online version contains supplementary material available at 10.1007/s13205-021-02828-2.

Keywords: Correlation; Enzyme-linked immunosorbent assay (ELISA); Real-time PCR (qPCR); Transgenic cotton.

Fig. 1

a Coomassie stained image of rCry1Ac migrating above 72 kDa. b Western blot image of rCry1Ac protein (above 72 kDa) c transgenic cotton (Bollgard II^®) leaf extract (~ 130 kDa) developed with SuperSignal™ West Pico Chemiluminescent Substrate

Fig. 2

mRNA relative quantitation versus protein wet weight for cry1Ac in leaves classified as a young and mature and b month of harvesting as one and three months. Regression lines for each category is shown as guidance

Fig. 3

mRNA relative quantitation versus protein wet weight for cry1Ac in a leaves, b squares and c stem. Blue line represents the best-fit regression and shaded region shows 95% confidence interval

Fig. 4

mRNA relative quantitation versus protein wet weight for cry1Ac in large, medium and solid squares. Regression line for each category is shown as guidance