OBJECTIVETo transform the antimicrobial peptide fusion gene of cecropin B and rabbit NP-1(CN) into Houttuynia cordata to improve its antimicrobic capability.

METHODThe fusion gene of CN designed and synthesized artificially was recombined with expression vector pBI121. The recombined vector was transformed to Agrobacterium tumefaciens LBA4404, by which CN gene was transformed to the explants of H. cordata. The transgenic regeneration plantlets were selected by kanamycin and rapid screening PCR. The transgenic plants were identified by PCR-Southern of genomic DNA and RT-PCR. The disease resistances were detected by antibacterial zone trail of leaf extracts to E. coli K12 and infection by Rhizoctonia solani.

RESULTGene of interesting CN was inserted into genomic DNA and expressed in transformed H, cordata, whose resistance to E. coli K12 and Rh. solani was stronger than that of the non-transformed control.

CONCLUSIONThe fusion gene CN can improve antimicrobic capability of transformed H. cordata.

Animals ; Anti-Bacterial Agents ; immunology ; pharmacology ; C-Reactive Protein ; genetics ; metabolism ; pharmacology ; Houttuynia ; genetics ; immunology ; microbiology ; Immunity, Innate ; Insect Proteins ; genetics ; immunology ; pharmacology ; Nerve Tissue Proteins ; genetics ; metabolism ; pharmacology ; Plant Diseases ; immunology ; microbiology ; Plants, Genetically Modified ; genetics ; immunology ; microbiology ; Rabbits ; Recombinant Fusion Proteins ; genetics ; immunology ; pharmacology ; Rhizoctonia ; physiology ; Transformation, Genetic

Carotenoid cleavage dioxygenase (CCD) family is important for production of volatile aromatic compounds and synthesis of plant hormones. To explore the biological functions and gene expression patterns of CsCCD gene family in tea plant, genome-wide identification of CsCCD gene family was performed. The gene structures, conserved motifs, chromosome locations, protein physicochemical properties, evolutionary characteristics, interaction network and cis-acting regulatory elements were predicted and analyzed. Real time-quantitative reverse transcription PCR (RT-qPCR) was used to detect the relative expression level of CsCCD gene family members under different leaf positions and light treatments during processing. A total of 11 CsCCD gene family members, each containing exons ranging from 1 to 11 and introns ranging from 0 to 10, were identified. The average number of amino acids and molecular weight were 519 aa and 57 643.35 Da, respectively. Phylogenetic analysis showed the CsCCD gene family was clustered into 5 major groups (CCD1, CCD4, CCD7, CCD8 and NCED). The CsCCD gene family mainly contained stress response elements, hormone response elements, light response elements and multi-factor response elements, and light response elements was the most abundant (142 elements). Expression analysis showed that the expression levels of CsCCD1 and CsCCD4 in elder leaves were higher than those in younger leaves and stems. With the increase of turning over times, the expression levels of CsCCD1 and CsCCD4 decreased, while supplementary LED light strongly promoted their expression levels in the early stage. The expression level of NCED in younger leaves was higher than that in elder leaves and stems on average, and the expression trend varied in the process of turning over. NCED3 first increased and then decreased, with an expression level 15 times higher than that in fresh leaves. In the late stage of turning over, supplementary LED light significantly promoted its gene expression. In conclusion, CsCCD gene family member expressions were regulated by mechanical force and light. These understandings may help to optimize tea processing techniques and improve tea quality.
Camellia sinensis/genetics* ; Gene Expression Regulation, Plant ; Phylogeny ; Plant Leaves/genetics* ; Plant Proteins/metabolism* ; Tea