2012 1839 tions through the means of ATP de- Dunn and others【8J used RNAi pin on actin and cysteine protease of ane- mone.proved inhibition of genetic ex— hybridization,and thus studied the functions of this gene. functions of special cancer genes and modifier genes,evaluating conven- tionaI cancer treatment and new pendence,and degrade RNA into dsRNAs of 19 to 21 bp.3’end of each cines,identifying growth marker ession of both genes through in situ medisection having 2 bases projecting.At prthe stage of RNAj reaction,two of tumor。analyzing the influence of tumor microenvironment upon the for— mation of tumor,and defining incuba- strands of siRNA bind with a ribozyme complex.and thus produces RNA-in— Study on signal transduction path- RNAi technology can easily confirm upstream and downstream duced silencing complex(RISC)问.In way order to activate RISC,there goes a process that ATP unwinds two strands of smalI molecule RNA.The activated tion and development stages of tumor. At present.mouse models of mamma- sequences of relative genes in the complicate signal transduction path- ways and their functions.James and others[ ̄;applied RNAi to study the sig- ry gland,pancreas,intestines,colon and prostate cancers have already been constructed,while models of RISC is Iocated at homologous mRNA transcript through base pairing,and brain,ovarium,neck,and skin cancers cuts mRNA 12 bases away from 3’ end of siRNA【 .Although the exact cu ̄ing mechanism is not clear,each RISC Includes a siRNA and a RNA enzyme different fr0m Dicer.Besides. there are also researches that show dsRNA with promoter region is also cut into sections 2 1-23nt long within plants.This dsRNA can lead to methylation of corresponding endoge— nous DNA sequences,and then pro— moters Iose functions and downstream genes silence. Application Status 0f RNAi Tech nology In recent years.as a highly effec— tive and specific genetic Interference technolgoy.RNAi develops very quick— IY and obtains great achievements of studies in basic biology,medicine, pharmacy,botany and other fields. Study of RNA-_n basic studies Study of genetic functions in eu- karyotic cells Along with the contin— UOUS development of genetic sequenc- ing technology and bioinformatics,ge- nomic sequencing of fruit flies,nema— todes.rice.rats and humans has al— ready finished,while the studies of ge- netic functions are relatively slow.I n comparison with traditionaI research methods of genetic functions finclud- ing gene knockout and antisense RNA).RNAi is a highly effective.spe— cific。quick and cheap method.1n combination with high-flux gene screening technologies as gene chip,it will play an important role in the exten— sive studies of genetic functions jn eu— karyotic cells.With the help of RNAi technology,genetic expression can be partly inhibited oR the leveI of RNA. achieving functional Ioss and further studying functions of relative genes. naI transduction pathway of insulin in are just at the beginning in studies fruit fly cell lines and obtained the re— RNAi js also used to construct suits totally same with known signal and protect other models of diseases transduction pathway of insulin.Be— along with construction of models sides,RNAi phenotype screening with— of cancerous diseases.Shcherbata[ jn the range of genome has already used heredity and RNAi to induce ac・ been an effective method In studying tivation of fruit fly cells and dystrophy the roles of genes in particular celI phenotypes as Iack of elasticity,mus- functions and biological activity,while cle degeneration dependent upon age, jt can not clearly distinguish the rela— and defective light sensing navigator. tionship among relative genes. Kim constructed slow virus of recom- Other fundamentaI studies ln addi— binants as transferring shRNAs into tion to the above two major applica, consemative sequence of Coxsackie tions,RNAi technology is also applied intestinal virus for the purpose of pre- to other fundamentaJ studies including venting viraI myocarditis and raising physiology and study into develop— the suwival rate of animaI models frOm ment and growth.PaImer and otherst viral myocarditisi .However.features used RNAi technology in the studies of animal models of diseases con— and application of physiology of ion structed through RNAi can neither Iast pathways in animal cells. very jong nor be jnherited steadily. Study on RNA..n medicine Furthermore,other problems still need Constructing modeI of diseases by to be studied. RNAi The traditiona1 way of con. Antivirus treatment through RNAi structing animal modeI of human dis— At present,studies of RNAi on the as. eases is gene knockout and Cre-LoxP pect of antivirus are basically to design systems.However.if the target genes cOrresp0ndIng siRNA to induce the are necessary to the growth of mice, degradation of specific mRNA and these methods are greatly restricted. thus be effective in antivirus.Novina of RNAi wilI be a new tooljn constructing MlT first used RNAi to resist HlV.fol— transgenic animaI mode1.which wilIln. Iowed by other cases of treating viraI activate gene functions specifying tis. diseases with RNAi。Afterwards,other sues along with its development.1t wilI scholars also took part in this research no Ionger need specifically conditioned as they designed various sirNA aga— mice and mice of tissue・specific re-・ inst diferent stages in HIV Iire cycle combination,and therefore transgenic and siRNA against genes gag,tat,reV, animals would be more widely used as nef and Olhers in HIV can be used t0 tools in biological studiesp1]. cOntrOI viraI reDlicatiOn【’4】.It has been Mice of genetic engineering have prOved that siRNA againSt sectiOn qa口 already been widely used to construct in viraI genOme can effectiveIV “si— modeI of human cancers and mice in lence”viraI genes and inhibit repliCa— the model simulate c0rreSpOndinq tiOn Of HIV virus【’3,’ ,’6】.Besides.there morphology,hist0pathOIOgy,pheno- were alsO Other sch0IarS Car ried Oul type and genotype of human cancers. studies jntO the functi0ns Of RNAi This kind modeI of diseases is better against Other diseases incIuding respi- than the traditionaI transgenic modeI of ratOry syncytiaI virus,SARS and pO- diseases in terms of analyzing the liOvirus.FOr exampI treatment Of hu. VOJ,1 3,NO,9,2O1 2 Agricu Jtura J Science&Techn0 JOgy 184O 2012 man and mice cells by siRNA of po. inhibited the expression of gene Alnylam Pharmaceuticals,whose RNAi Iiovirus genome can noticeably reduce MTA1 in MDA-MB.231 and MCF.7 by medicines were to cure respiratory the amount of virus in filiaI genera. RNAi in order to lower expression of syncytial virus,influenza,Parkinson tions,and speed up the elimination of ERc(。and then invasion activity of disease and hypercholesteremia.The virus frOm infected cells]切. breast carcinoma cells in both plants other was Sirna Therapeutics,and Study on RNAl in tumor treatment decreased. their RNAi medicine to treat ADM.Sir. As RNAi technology can effectively si. Moreover.Wang and others【筮】 na-027.has already been in clinical Ience specific target genes.1t becomes designed RNAi against most powerful Phase 1.Other applications as curing a power research tooIin determining survivin up to date.and transferred in- Huntington,asthma and COPD are al- the role of a single gene jn tumor for. tO KYSE51 0.carcinoma celI Iine of so being studied before clinical appli- mation and development and explor— human esophageal squamous epithe・ cation.However.there are stilI prob- ing genetic treatment of tumor.Natu- liaI cells.They proved thal down regu- lems with RNAi medicines in terms of rally it has a wide application. Iation of surviving could strongly inhibit supply source and safety in usage. Activation of proto oncogene and/ the growth of internal and in vitro can. fInformation is fr0m h ̄pJ/www.gan- or inactivation of anti—oncogene will re— cer cells and presumed this inhibition baobao.com.cn m怊 一339333.htmI.1 suit in tumor.Since RNAi technology could promote the increase of apopto- Study on RNA_.n botany enjoys an advantage in the study of sis.This study would deepen the RNAi phenomenon was first single genes and RNAi can inhibit knowledge about pathOqenesis of eso- found in plant study and therefore.ap— cancer genes in tumor and functionaI phageal carcinoma,explain that sur・ plication and study of RNAi technology expression of anti—oncogene,analysis vivin could possibly be the potential is relatively thorough in plant。including of its phenotype changes can help re— target site for treating esophageal car・ studies of plant functionaI genomes. veaI the molecular mechanism of tu. cinoma.and RNAi had a potentiality in growth and development,disease re・ mor formation and development and treating diseases.At present.in China sistance,and improvement[ ̄. COnsequently provide marker and tar- there have been researches into RNAi Study on plant functional genomes get for tumor diagnosis and treatmerit。 inhibition of survMn expression in Since post-genome era is on the Wang and others[18】used RNAl to in. pancreas cancer,cervical cancer,lung way,there is an increasing need for teffere with the expression of CML66 cancer,colorectaI carcinoma,neurob・ study into 9enetic functions in an ex- in celI Hela and then found its prolifera- Iastoma cancer,osteosarcoma cells, tensive and high—flux way.Due to the tion,invasion and metaptosis are in- and Ieukemia cells,yet most of them fact that RNAi is highly specific ln se- hibited.Further study on its mecha— are stilI at the stage of in vitro study. quence and effectively interferentia1.It nism indicated that after the inhibition Study on RNAi oR medicine can specifically silence certain genes of the expression of CML66,cathepsin RNAi is applied to medicine re— and thus obtain deficiency or reduce L,MMP1 5,uPAR,VEGF。COX-2, search as It can specifically and effec- mutation.Therefore.it can be used as S1 00-A4.MUC1.MDM2 and RAC1 a¨ tively inhibit genetic expression,obtain an effective toolin studying genomics。 decreased,and therefore,the conciu- afunctional phenotype,mass screen ln November.2002.the Ara— sion was as an oncogene.CMI 66 can target in a short time,and therefore, bidopsis genomic RNAi knock—out Iine promote proliferation,invasion and screen and assess drug targets on celI analysis (AGRlKOLA)ajready pre— metaptosis of tumor through various IeveI and animallevel[23].Hamamichi pared tO use RNAi technOIOgy lO study meaas. exploited RNAl to systematically shield the functions of aIl qenes (about Study on the application of RNAi nearly 900 candidate genes and path- 25000)of Ara6fd ps『s a/『ana.The technology to mechanism of tumor in— ways in model of Parkinson disease。 apprOach is tO use all genetic pieces vasion and metaptosis as welI as and found 2 genetically susceptible Of raDf2,DDs『S a/『a门a tO cOnstruct changes of relevant genes is good to sites and targets of Parkinson disease, cOnst_tutive and inducibIe RNAi car rier fully understand the biotogical nature which became the foundation for the and transfer ra6/db()s,s a『『ana of malignant tumor.Shin and others㈣ treatment of this disease. 廿1rOuah aarObacte rium mediatjOn tO used RNAj technology to knock out While RNAi is widely applied to aet mOre than 4 O00 COnstitutlve hp— expression of eaveolin-1 in order to study of functional genome to spot RNA plants【 .MOreOver,Travella and regulate the expression of vascular many potential targets of medicines,it OtherS嘲suCcessfuIly inhibit target gen— celI adhesion molecule、1(VCAM-1) is also proved to be a new possible es in the cOnstructed he×aDIOid wheat and cOnsequentlV adhesion of tumor medicine for gene therapy.1n 2006, transgenic crOps Of PDS—RNAi and cells dependent upon VCAM一1 Is sig- Bevasiranib.RNAi medicine manufac— EIN1.RNAi.Lawrence and Others[28】al_ nificantly inhibited.Tsuchiya and oth- tured by American Acuity Pharmaceu— sO used dsRNA reIevant tO methvIa- ers prepared I nhibitor of DNA bind- ticals to treat humid senile plaque tiOn Of ,a6fdbDs『s a/7al仃a cen— ing proteins 1,3(Idl and Id3)by RNAi pathology.was successfuI in clinical lr0mere fOr RNAi Of raD s,S to double knock out gastric cancer celI trials since it was better than conven— 1a/ i门a tetrapIOid. They fOUnd the Iine MKN45 and proved that knockout tionaI medicines in lts effect and safe・ c0ntent 0f mef}1vIatiOn-reIate耍 RNA Of Idl and ld3 could significantly inhibit tv.1n addition to Acuity,two other in A,_abfdOpsjSl fna』fana tetraDIOjd de- metaptosis of peritoneum of gastric companies also took part in the devel— creased sharpIV. All廿1e abOve can cancer cells.And Jiang and others[21 】opment of RNAi medicines.ORe was demOnStrate RI、JAi teChnOIOgy iS a Agricultural Science&Technology Vo1.1 3,No.9,201 2 2012 good tooIin deficiency and mutation of polyploid plants. Study on plant growth,develop- ment and breeding new varieties Male sterility is common in higher key enzyme in synthesis of flower co1. or and flavonoid,and turned the co1. Or frOm originaI blue into white and existing Iife phenomenon in nature, and its technology wilI definitely play an important role in Iife sciences.and therefore.its application will have a pink,producing a new commercial flower.Australian and Japanese re— searchers eliminated gene DRF in great future. plants and has enormous value in agriculture.Cigan and others i291 used rose to produce the unique blue rose jn the world【3引.AI lof these show lhe particular economic value of RNAi technology jn regulating flowers’or. namentaI organs including color. References [1 1 MATZKE MA,PRlMIG M,TRNOVSKY J.ef a1.Reversible methylation and in. activation of marker genes in sequentia1. RNAi technology to straightly silence M¥45,promoter for genetic expres— sion of maize pollen sac.and then in— hibit the formation of pollen sac to produce maize male sterile plant. RNAi technology can produce some 1y transformed tobacco plants[J].EMBO, 1 989.8:643-649. Study on plant resistance Resis. tance mechanism mediated by RNA f2】NAPOLl C,LEMIEUX C,JORGENSON perfect sterile materials.Moritoh and others[3o}obsewed OSGEN-L-RNA rice plant and found some with low fertility, some infertility.becoming the male sterile materials of rice. 1ncrease or decrease of the active elements in economic crops can in— crease their economic value and uti— lization rate.Zhang Yinbo and othersp1】 cloned PEP pieces of rape bv RT.PCT technology and constructed RNAi car. rier cOrresDOndent to gene PEPASE in order to inhibit genetic expression of PEPASE through RNAi technology. Therefore,metabolism flow would be more of oil synthesis and the oiI con— tent in rape seed would increase.Fur— thermore,specific promoter of fruit was also used with RNAi technology to inhibit morphology of tomato endoge- nous Iight and construct expressions regulating gene DET.Finally,expres‘ sion of DET1 in regenerated plants re— duced and the content of carotenoid and flavonoid increased noticeably. while other parameters of fruit did not change greatly.All these indicate that silencing of specific genes in organs can improve the nutrition value of plants.Also.Shinjiro and others In- hibit genes controlling caffeine syn- thesis with the help of RNAi technolo- aV and consequently the content of caffeine in the materials reduced 50% to 70%. RNAi technology can make some improvements for certain plant phe- notypes.which provides much fun for people’s Iife and increases more eco— nomic value as wel1.To a certain de- gree,it also increases plant varieties, especially improvement for flower quality.With the help of RNAi technol— ogy,Japanese Biological Research lnstitute successfully Induce the si- lencing of chalcone synthase(CHS),a used dsRNA expressing virus source. which is an effective method to get re. sistant transgenic plants at present. Many scholars are trying to conduct studies into plant virus through this technology,while the most used is transformation mediated by agrobac. terium integrate s dsRNA into plant genomes,and then test resistance of transformant against virus.or after in- stant transformation of the system by agr-obacterium test plants for viraI in- fection during instant expression of dsRNA in order to determine effective- ness of viral’resistance mediated by dsRNA, Up to date,there have been many reports about the application of RNAi technology to plant virus prevention, yet the lechnology is not quite mature. First,not alI RNA sequences are that easy to be recognized and cut by siR- NA because some probably hide in great folds so that they are not recog- nized by siRNA.Second.mutation is prone to happen during virus replica。 tion.and hence antibodies faiI to func. tion for a Iong time.There are also problems conceming hereditary stabili- tV to be solved.Therefore.more thor- ough and detailed studies are needed in the respect of prevent of plant virus with RNAi. Conclusion In a word.RNAi not only greatly promotes the development of post-hu- man genome project(proteomics),but also has profound influences on the aspects of gene therapy,development of new medicines,improving plant breed.increasing plant variety and promoting plant economic value.AI— though the present study of RNAi is not thorough enough and RNAi tech. nology is not perfect.RNAi is a widely VOI.1 3,NO.9,201 2 R.Introduction of a chimeric chalcone synthase gene in petunia results in re— versible co—suppression of homologous genes in trans[J].Plant Cell,1 990,2(4): 279-289. [3】GUO S,KEMPHUES KJ.Par-1,a gene required for establishing polariry in C. elegans emb ̄os,encodes a putative Ser/Thr kinase that is asymmetrically distributed[J].Cell,1 995,81(4):61 1_620. 『41 FIRE A,XU S,M0NTGoMERY MK.ef a1.Potent and specific genetic interfer. ence by double—stranded RNA in Caeno- rhabditis elegans[J].Nature.1 998,391: 806. 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Down regulation of surviving by RNAi inhibits the grown of esophageal carci- tion in plants(RNAi技术的研究及其在 植物上的应用)fJ】.Journal of Anhui A. crospore development in rice『J1.Plant CelI Physiology.2005.46:699—71 5. gricultural Sciences(安徽农业科学), 201 0,38(8):3897-3899. noma Cells[J].Cancer Biol Ther,2005, 【31】ZHANG YB(张银波),JIANG ML(江木 4(9):974-978. 兰),HU XJ(胡小加).Rapeseed Responsible editor:Yingke CHEN Responsible proofreader:Xiaoyan WU I ̄NAi的研究进展 张俐 ,张雅琼 ,余丽娜 ,徐文婷3贾阳映 ,谢世清 ,孙文丽 ,梁泉 (1.云南省科学技术情报研究院,云南昆明650051;2.云南中医学院, 云南昆明650500;3.云南农业大学农学与生物技术学院,云南昆明650201) 摘要RNAi是由与靶基因同源的内源或外源双链RNA(double stranded RNA,dsRNA)所引发的一种在动植物中普遍存在的基因沉默现象。它 最早在植物体中被发现,现在已发展成为一种生物技术,并成为了后基因时代的重要研究手段。对RNAi技术在生物基础、医学、药学、植物学等 领域的研究成果进行了综述。 关键词RNAi;后基因组;功能基因组 作者简介 张俐(1978一),女,云南保山人,硕士,助理研究员,研究方向:科技查新及项目申请管理。张雅琼(1979一),女,山西太原人,博士,从事 药用植物资源评价与利用方面的研究。 共同第一作者。 通讯作者,高级农艺师,从事植物营养与分子育种研究,E.mail: liangquan1@l63.com。 收稿日期 2012-01-06 修回日期2012—08—16 Agricultural Science&Technology Vo1.1 3,No.9,201 2