In-planta transient transformation of avocado (Persea americana) by vacuum agroinfiltration of aerial plant parts
In-planta transient transformation of avocado (Persea americana) by vacuum agroinfiltration of aerial plant parts
复制标题
DOI：
10.1007/s11240-022-02436-9
复制
发表时间：
2023-01-15
影响因子：
3
通讯作者：
Urrea-Lopez，Rafael
中科院分区：
文献类型：
--
作者：
Salazar-Gonzalez，Jorge Alberto;Castro-Medina，Manuela;Urrea-Lopez，Rafael
鳄梨是国际市场上交易量最大的热带水果之一。到目前为止，只有合子胚胎实现了稳定和瞬时的转化，而成年植物组织则没有实现转化，这限制了功能基因组学的研究。我们提供了第一个瞬时农杆菌介导的鳄梨叶片转化方法，克服了该物种对转化的抵抗。研究了农杆菌、叶期、伤害前处理、植物激素茉莉酸和真空渗入对鳄梨叶片瞬时转化的影响。利用农杆菌LBA4404和RUBY对牛油果离体叶进行转化，转化率可达27%。转化效率与叶龄有关，叶片发育中期的瞬时报告基因表达效率最高。微伤前处理促进了土壤入渗，叶龄是影响瞬时转化能力的主要因素。在没有微损伤的情况下，茉莉酸对瞬时转化没有显著影响。然而，微创和250微米茉莉酸协同作用显著提高了瞬时表达。利用这种局部真空农业渗透的方法，实现了牛油果叶片的瞬时转化。这种方法开启了利用农杆菌介导的瞬时转化作为一种工具来探索鳄梨叶片的基因功能和代谢途径的方法。
Avocado,Persea americanaMill, is one of the most traded tropical fruits in the international market. To date, stable and transient transformation has only been achieved for of zygotic embryos and not of adult plant tissue, which limits functional genomics research. We provide the first transientAgrobacterium-mediated transformation methodology in avocado leaves that overcomes the recalcitrance to transformation of this species. We investigated the effect ofAgrobacteriumstrain, leaf stage, wounding pre-treatment, the phytohormone jasmonic acid, and vacuum infiltration on transient transformation of avocado leaves. Using theAgrobacteriumstrain LBA4404 and theRUBYreporter a transformation frequency of up to 27% was obtained for avocado detached leaves. The transformation efficiency depended on the age of the leaf, with an intermediate stage of leaf development showing the highest efficiency of transient reporter gene expression. Microwounding pre-treatment facilitates agroinfiltration and coupled with leaf age are the primary factors influencing competence for transient transformation. Jasmonic acid did not significantly affect transient transformation in the absence of microwounding. However, microwounding and 250 µM of jasmonic acid acted synergistically to significantly enhance transient expression. Using this methodology with localized vacuum agroinfiltration, transient transformation of attached avocado leaves was achieved. This method unlocks the use ofAgrobacterium-mediated transient transformation as a tool for explore gene function and metabolic pathways in both, detached and attached avocado leaves.