Construction of Cinnamomun bodinieri cDNA library and screening of upstream regulators of CbP5CS using yeast one hybrid

Abstract Alkaline soil is a main factor limiting the introduction and popularization of Cinnamomun bodinieri. Proline is one of the most important compatible solutes that plants accumulate for osmotic adjustment in response to alkali stress. Therefore, it is of great significance to identity the key transcription factors that regulate the biosynthesis of proline and illustrate the underlying molecular mechanisms. In the present study, we constructed a C. bodinieri cDNA library using alkali treated root tissues. The titer of the library is 4.88×107CFU/mL. The average size of the inserts is 1000 bp, and the recombination efficiency is 100%. Then we cloned the promoter region of CbP5CS and screened the CbP5CS promoter interacting proteins by yeast one hybrid and high-throughput sequencing. Our results showed that 31 unique CbP5CS-interatcting ESTs were identified and 6 of them were annotated as transcription factors, including GW020491 (zinc finger CCCH domain-containing protein 25 isoform X1), GW028183 (AtbHLH104), GW000650 (transcription factor TFIIIC), GW007525 (RING/FYVE/PHD zinc finger superfamily protein), GW015686 (GATA type zinc finger transcription factor family protein), GW027120 (AtbHLH96). These transcription factors may play a key role in regulating the production of proline.