Abstract
Yeast two-hybrid and one-hybrid technologies are an efficient molecular biology technique for screening protein interactions or protein and DNA interactions, and are an important means for the interaction and regulation of biological macromolecules. In order to obtain a high-capacity Medicago truncatula gene library, it provides a basis for further digging the related genes of Medicago truncatula and improving the quality of Medicago truncatula . In this study, Medicago truncatula from different tissue sources was selected, and different hormone induction or stress treatments were performed on Medicago truncatula , using SMART technology to successfully construct a high-capacity Medicago truncatula yeast hybrid cDNA library. The library quality test results showed that the library titration number was 5×10 7 CFU/mL, the library capacity was 1.28×10 7 CFU, and the average insert fragment length was greater than 1 000 bp. All 24 clones were able to amplify bands, and the cDNA fragment recombination rate was 100%. The library has high quality and complete genetic information, which meets the requirements of yeast hybrid screening test, and can be applied to the research of gene expression regulation of Medicago truncatula and the screening test of interaction protein.
Highlights
Plant cDNA library refers to the clone set formed by the reverse transcription of all mRNAs of plant tissues at a specific growth and development period into cDNAs, and connecting them with appropriate library vectors and transferring them into specific bacteria (Gubler and Hoffman, 1983)
Medicago truncatula from different tissue sources was selected, and different hormone induction or stress treatments were performed on Medicago truncatula, using SMART technology to successfully construct a high-capacity Medicago truncatula yeast hybrid cDNA library
1.1 Good mRNA isolation quality of Medicago truncatula After 1% agarose gel electrophoresis detection (Figure 1), the results showed that the purified mRNA was a fuzzy drag band, and the brightest part of the drag band was more than 500 bp, indicating that the isolated and purified mRNA did not decompose and had good quality, which could be used as the initial sample of the library
Summary
Plant cDNA library refers to the clone set formed by the reverse transcription of all mRNAs of plant tissues at a specific growth and development period into cDNAs, and connecting them with appropriate library vectors and transferring them into specific bacteria (Gubler and Hoffman, 1983). Plant cDNA library can be used to screen regulatory genes in plant growth and development stages, and play an important role in the prediction of interaction proteins and in vitro expression (Wang et al, 2014). Plant cDNA library plays an important role in discovering and studying new gene functions, and is favored by researchers (Yang et al, 2007). Compared with the related species of Medicago sativa and other plants in Leguminosae, Medicago truncatula has many genetic advantages, such as small genome, high efficiency of genetic transformation, short life cycle, self-pollination and so on (Blondon et al, 1994; Trieu et al, 1996; Bonnin et al, 1997), which make it a model plant of Leguminosae. Legume Genomics and Genetics 2022, Vol., No.1, 1-7 http://cropscipublisher.com/index.php/lgg can be used for screening protein interactions or protein and DNA interactions in various mechanisms, which provides a basis for further research on basic biology such as protein interaction or protein and DNA interactions
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