The Genetic Transformation Of Lotus Corniculatus L. By Antisense 4CL Gene And LePT1 Gene

Lotus corniculatus L.,also known as marsh birdsfoot trefoil,is found worldwide.There is a flourishing demand for Lotus corniculatus L.as an important protein sourc e fodder,leguminous green manure and grass for pasture afforestation and improveme nt in the animal husbandry and grass industry.Lignin is one of the major components of cell walls.Rich lignin content and stab le structure make leguminous forages less biodegradable within a short time and beco me an impediment to the forage digestion and absorption by livestock,and thereby th e utilization of forages can be improved by reducing lignin content.4-coumarate:CoA ligase(4CL)locates in the branch points of synthesis of phenyl methane derivatives and is a key enzyme in lignin biosynthetic pathway.Research has shown that lignin c ontent can be lowered by inhibiting 4CL activity.Phosphorus is one of the essential nutrients needed for plant growth.The phosp horus-deficient areas cover around up to 40% of the arable soil in the world,which pr events the proper growth of crops.Phosphate fertilizer usage is increasing,but it sees a low usage in season.Phosphate fertilizers are vulnerable to being cured by arable la nd,causing the pollution of phosphorus concentration in soil and the approaching dep letion of phosphorous metal resources.Therefore,it is of great significance to increas e the absorption and utilization of soil phosphorus by plants.LePT1 gene,an efficient phosphate transporter gene cloned from tomato genome and responsible for regulatin g the transport and absorption of phosphorus by plant roots,is a high-affinity phospha te transporter family under the most intensified research.Plant genetic transformation is an effective breeding path to improve crop traits.This research focused on cotyledonary nodes of Lotus corniculatus L.as explant materials to explore the effects of the hormones of different kinds and concentrations on callus i nduction,proliferation and differentiation and rooting of Lotus corniculatus L.A set o f high frequency regeneration systems for tissue culture of Lotus corniculatus L.was established,laying a foundation for the follow-up genetic engineering experiments ofLotus corniculatus L.By using the above-mentioned high frequency regeneration syst ems for tissue culture of Lotus corniculatus L.with mainly the procedures like agroba cterium infection co-cultivation,recovery culture,differentiated screening culture and rooting culture,the T0-generation transformed lotus corniculatus was produced.By means of PCR tests and target fragment sequencing verification,1 positive 4CL trans genic plant and 5 positive LePT1 transgenic plants were produced.After T0 generatio n plants were transplanted and survived,they flowered and seeded under conventional management.This has made a contribution to breeding new lotus corniculatus strain with low lignin content and the highly-efficient utilization of leguminous forage root system to phosphorus under deficient phosphorus stress environment.What's more,it has laid a solid foundation for researches into genetic transformation of leguminous f orages.