继代培养次数对成年鳄梨茎段生长发育及复幼效果的影响OA

Avocado,a fruit of significant economic and nutritional value,still faces technical challenges in tissue culture,with explant maturity being a key factor influencing cultivation success.This study utilized new shoot segments from mature avocado trees as explants.By establishing culture experiments with different subculture times,the effect of subculturing on shoot rejuvenation was investigated,aiming to provide a technical basis for improving avocado micropropagation efficiency.Growth indicators of new shoots(including length,stem diameter,leaf number,and leaf area)were systematically measured,and changes in the relative expression levels of miRNA156 and miRNA172 were analyzed to assess the correlation with the rejuvenation state.Results showed that with increasing subculture frequency,new shoot length,stem diameter,leaf number,and leaf area all increased significantly,indicating that a higher subcul-ture number promotes shoot morphogenesis and leaf expansion.At the molecular level,after the second subculture,the ratio of miRNA156/miRNA172 relative expression in new shoots reached its highest value(2.32),marking the peak of rejuvenation.Subsequently,the relative expression of miRNA156 gradually declined,suggesting a transition of the shoots towards maturity.By the sixth subculture,the expression level of miRNA172 exceeded that of miRNA156,further indicating the shoots entered a mature state.Morphological and anatomical observations revealed that from the fifth subculture onward,lignification of the stem segments intensified,and xylem structure became more complete.Simulta-neously,starting from the fifth subculture,pale yellow granular callus gradually appeared at the basal cut ends of stem segments.The amount and volume of this callus increased noticeably with further subculturing(up to the seventh sub-culture).Although root induction was not successful in any treatment,the observed morphological changes suggest that the rooting potential of the material may be gradually enhanced with increased subculture frequency.In conclusion,continuous subculturing can effectively promote the rejuvenation of mature avocado rootstock material,leading to ob-vious juvenile characteristics.Based on growth indicators,miRNA expression patterns,and morphological observations,it is recommended to control the number of subcultures within five for optimal rejuvenation effects.These findings pro-vide experimental evidence and a reference for rejuvenation techniques in the tissue culture of mature avocado plants.