Preliminary Study On In Vitro Culture From Persea Americana Mill ‘Hass’

Avocado（Persea Americana.Mill）being to the family Lauraceae,is cultivated in tropical and subtropical areas worldwidewith rich nutrition value.An efficient and stable regeneration system is a pre-requisite for plant genetic transformation and the cultivate for new varieties.Avocado is difficult to regenerate from in vitro-cultured explants,which has largely hindered the development of new variety using the tools of modern biotechnology.With in vitro tissue culture,avocado can be in rapid multiplication without seasonal restriction.In this study,using segment,leaf,petiole and stem apex from avocado‘Hass’as the explants and effect of basal medium,plant growth regulator types and content and types of explant on adventitious shoot regeneration and callus induction culture were investigated.This study preliminary established an in vitro regeneration system from avocado‘Hass’,which laid the foundation for further in vitro culture system for avocado‘Hass’and others.Main experimental results were as follows:Establishment of regeneration system with callus:（1）Feasible disinfection and sterilization method:The explants（leaf,petiole and stem）were collected from March to August,which were surface sterilized with 75%（w/v）ethanol for 20³0 s,followed by washed with sterilized distilled water for two times,followed by 5⁶ mins in 0.1%HgCl₂（m/v）,finally washed thoroughly in sterile distilled water for 4⁵ times;（2）The optimun medium for callus induction was:MS+1.0 mg·L^-1 6-BA+1.0mg·L^-1 NAA+30 g·L^-1 sucrose+7.0 g·L^-1 agar,pH=5.8±0.1（cultured under dark environment,84%）;（3）The best kind of explants for callus induction was segment,petiole followed by,leaf and stem apex were the worest;（4）The optimun kind of basal medium for callus induction culture was H basal medium（72.1%）,N₆ and MS followed by（61.4%and 39.5%respectively）,WPM,B_5modiedmodied and 1/2MSwere the worest（respectively 27.6%,16.2%and 11.6%）;（5）Adventitious bud induced directly from leaf explants:under the light environment,on the H induction medium contained 6-BA and NAA,there was no adventitious shoot buds developed directly on leaf explants but coubld be induced callus;（6）Adventitious bud induced from callus:there was no bud differentiation from callus in the differentiated medium supplemented with NAA and 6-BA;（7）Callus induced from ather explants:it was difficult to induce callus from anther explants of avocado‘Hass’.Establishment ofregeneration system with nodal segments（semi lignifications）:（1）Suitable disinfection and sterilization method:The stems explants were collected from March to August,which were surface sterilized with 75%（w/v）ethanol for 20³0s,followed by washed with sterilized distilled water twice,followed by 6⁸ mins in 0.1%HgCl₂（m/v）,finally washed thoroughly in sterile distilled water for 4⁵ times;（2）The best kind of basal medium for bud induction was MS（73.7%）,H followed by（66.6%）;（3）The optimun medium for bud induction was:MS+1.0 mg·L^-1 6-BA+0.2mg·L^-1 IBA+30 g·L^-1 sucrose+7.0 g·L^-1 agar,pH=5.8±0.1（82.5%）;（4）The optimun medium for shoot multiplication was:MS+1.0 mg·L^-16-BA+0.2 mg·L^-1 NAA+30 g·L^-1 sucrose+7.0 g·L^-1 agar,pH=5.8±0.1.（5）It was hard to induce rooting with in vitro plantlets for avocado’Hass’,it is needed to make thorough research.