纳米磁珠介导牡丹花粉转基因技术的初步分析

Preliminary Analysis on Transgenic Technology of Pollen-Mediated by Magnetic Nanoparticles in Tree Peony

磁性纳米颗粒(magnetic nanoparticles,MNPs)可以吸附DNA,作为一种新型基因载体应用于转基因育种。本研究旨在探索MNPs介导外源基因进入牡丹花粉,为利用携有外源基因的花粉进行授粉,获得转基因种子提供依据,本研究制备了MNPs与DNA的复合物;利用扫描电子显微镜与透射电子显微镜观察了牡丹花粉的形态结构;用花粉离体萌发法测定了供试牡丹花粉磁转染前后的生活力。结果表明:MNPs与DNA的质量比为1:1~1:10时,MNPs均可以与DNA稳定结合,并可以保护DNA免受酶的消化作用;牡丹花粉为长球形,萌发孔为三孔沟类型,位于萌发沟内;‘凤丹’牡丹、‘京红’牡丹及紫斑牡丹的花粉均具有较高的萌发力,且经磁转染后的三种牡丹花粉萌发力保持在50%以上,能够用于正常杂交授粉。本研究首次采用四氧化三铁纳米磁转化技术介导牡丹花粉转基因,弥补了花粉介导牡丹转基因方法研究的空白,从而有助于牡丹分子育种进程的推进。

As a kind of new gene carriers,magnetic nanoparticles(MNPs)can be applied to transgenic breeding because it can absorb DNA.This study aimed to explore the technology by which the MNPs can deliver exogenous genes into tree peony pollens so that provide an reference for transgenic breeding of tree peony through pollination with pollens containing exogenous genes.In the study,MNPs/DNA complexes were prepared;the morphology of tree peony pollens was observed by the scanning electron and transmission electron microscopes;the germination of magnetofected and non-treated pollen grains in vitro were observed.The results revealed that MNPs have effectively been loaded with DNA when the mass ratio of MNPs to DNA was 1:1~1:10,which could also protect DNA against enzymatic degradation.The tree peony has nearly spheroidal,tricolporate pollens with the apertures that lay in the furrow.The pollen of Paeonia ostii'Fengdan'and P.rockii'Jinghong'with ideal germination viability could still be used for crossing whose germination viability maintained above 50%after magnetofection.In this study,we adopted magnetofection technique based on Fe3O4 nanoparticles to achieve pollen-mediated tree peony transformation for the first time,which made up for the blank of transgenic method of pollen-mediated tree peony,so as to promote the molecular breeding of tree peony.