Heterologous expression of Hp BHY and Cr BKT increases heat tolerance in Physcomitrella patens

Heat stress can restrict plant growth,development,and crop yield.As essential plant antioxidants,carotenoids play significant roles in plant stress resistance.b-carotene hydroxylase(BHY)and b-carotene ketolase(BKT),which catalyze the conversions of b-carotene to zeaxanthin and b-carotene to canthaxanthin,respectively,are key enzymes in the carotenoid biosynthetic pathway,but little is known about their potential functions in stress resistance.Here,we investigated the roles of b-carotene hydroxylase and b-carotene ketolase during heat stress in Physcomitrella patens through expressing a b-carotene ketolase gene from Chlamydomonas reinhardtii(Cr BKT)and a b-carotene hydroxylase gene from Haematococcus pluvialis(Hp BHY)in the moss P.patens.In transgenic moss expressing these genes,carotenoids content increased(especially lutein content),and heat stress tolerance increased,with reduced leafy tissue necrosis.To investigate the mechanism of this heat stress resistance,we measured various physiological indicators and found a lower malondialdehyde level,higher peroxidase and superoxide dismutase activities,and higher endogenous abscisic acid and salicylate content in the transgenic plants in response to high-temperature stress.These results demonstrate that Cr BKT and Hp BHY increase plant heat stress resistance through the antioxidant and damage repair metabolism,which is related to abscisic acid and salicylate signaling.

A case study of using an efficient CRISPR/Cas9 system to develop variegated lettuce

The clustered,regularly interspaced,short palindromic repeat associated endonuclease 9(CRISPR/Cas9)system has emerged as a powerful approach for precision breeding to create plants with desirable traits.However,the CRISPR/Cas9 system relies heavily on an efficient plant transformation system that is usually time-consuming and costly.Here,we have constructed a CRISPR-Cas9 vector with neomycin phosphotransferase II and green fluorescent protein(eGFP-NPTII),where the high expression of GFP during plant regeneration allowed us to minimize the positional effect on T-DNA expression and facilitate screening T-DNA-free mutants.Successful gene editing using CRISPR/Cas9 has been illustrated in different plant species,but an important aesthetic characteristic of leaf variegation remained unexplored.With the newly designed construct,we have targeted the variegation gene LsVAR2 in lettuce.Our results indicated that LsVAR2 is closely related to both AtFtsH2 and AtFtsH8,in which homozygous mutations lead to an albino phenotype while a variegated phenotype was induced by CRISPR/Cas9 de novo gene editing.In conclusion,the unique design of our CRISPR/Cas9 construct could efficiently edit the target gene and ease the screening of non-TDNA mutants through detecting GFP signals during plant regeneration and progeny segregation.Additionally,the success of gene-editing of LsVAR2 in lettuce demonstrates proof in this method to develop novel plant breeding materials for valuable horticultural plant species.