[生物工程精品] 粗毛栓菌漆酶cdna的pcr扩增 论文

本科生毕业设计（论文）题目粗毛栓菌漆酶CDNA的PCR扩增姓名学号系别生命科学系专业生物科学指导教师职称教授年月日教务处制A1A0A2A3A4A5A6A8A9A7A11A10A12A13A14A15A16A17A18A19A20A21A22A23A24A25A26A27A28A29A30A31A32A33A34A35A36A37A38A39A40A41A42A43A44A45A46A47A48A23A49A50A27A51A52A49A53A54A55A56A57A58A55A56A59A60A61A27A62A63A26A64A65A27A66A67A68A40A35A36A27A69A40A70A71A38A39A40A43A72A73A74A75A76A77A40A70A71A38A39A40A78A79A80A33A34A35A36A81A82A83A41A42A24A84A43A85A86A87A86A87A86A87A88A88A89A90A91A26A27A31A32A33A34A35A36A27A40A70A71A38A39A40A41A42ABSTRACTTOTALRNAWASEXTRACTEDFROMTRAMETESGALLICACULTIVATEDFOR10DAYSINHIGHCLOWNMEDIUMTHERESULTSHOWEDTHATTWOCDNAFRAGMENTSWASAMPLIFIEDBYRTPCRKEYWORDSTRAMETESGALLICAFR,LACCASECDNA,RTPCRA92A93A94A95A96A97A98A99A100A101A102A103A104A105A106A107A108A109A110A111A112A113A114A115A116A117A118A119A120A121A122A123A124A125A126A127A128A120A129A130A131A132A133A134A135A136A137A138A131A139A140A120A141A142A143A144A137A145A146A147A148A149A150A151A152A153A121A122A154A155A120A156A157A137A158A131A151A159A160A145A146A161A162A149A150A151A155A163A120A164A165A137A153A161A111A151A166A167A168A169A170A110A118A164A165A137A162A171A162A172A173A118A174A175A170A103A104A105A106A176A177A109A178A179A180A173A118A181A182A109A183A184A185A186A187A176A188A189A109A178A179A180A183A190A191A192A193A109A183A194A188A109A178A179A180A195A196A197A198A199A200A201A202A203A204A205A203A206A207A208A209A210A211A204A212A213A210A214A215A216A217A218A216A219A220A221A222A223A224A222A225A226A217A227A228A229A230A231A232A233A234A235A236A237A238A239A231A236A239A240A241A242A243A244A245A246A236A239A247A234A235A222A248A249A250A251A222A252A253A254A255A235A222A66A0A1A2A235A222A9A3A4A3A208A5A6A240A7A66A8A217A75A10A11A244A231A12A13A14A15A16A17A18A19A20A21A22目录摘要1关键词1ABSTRACT1KEYWORDS1引言11材料与方法211主要仪器与试剂用品2111主要仪器2112主要试剂2113主要用品212菌株及培养方式2121菌株2122培养基313PCR引物314总RNA的提取与检测3141总RNA的提取3142总RNA的甲醛变性琼脂糖凝胶电泳4143总RNA纯度与浓度的检测415RTPCR4151逆转录4152总CDNA的PCR扩增4153PCR扩增产物的琼脂糖凝胶电泳52结果与讨论521总RNA电泳结果522总RNA纯度与浓度523PCR扩增结果6参考文献6致谢7A23A24A25A26A27A28A29A30A31A32A33粗毛栓菌漆酶CDNA的PCR扩增生物科学专业学生于林田指导教师孙迅摘要本实验以粗毛栓菌作为出发菌，进行了漆酶CDNA的PCR扩增。以半纤G13512G13044为G11911G9316，用G20652G11911G1314G8706培养基G15培养1G19D，G6922G19610菌G1009G1319，提取总RNA，G1582RTPCR。结果G7186G12046G17902G17819RTPCRG7389G1016G7477CDNAG10267G8585G15999扩增出G7481。关键词粗毛栓菌，漆酶CDNA，RTPCR扩增PCRAMPLIFICATIONOFLACCASECDNAINTRAMETESECGALLICASTUDENTMAJORINGINBIOLOGYYULINTIANTUTORSUNXUNABSTRACTTOTALRNAWASEXTRACTEDFROMTRAMETESGALLICACULTIVATEDFOR10DAYSINHIGHCLOWNMEDIUMTHERESULTSHOWEDTHATTWOCDNAFRAGMENTSWASAMPLIFIEDBYRTPCRKEYWORDSTRAMETESGALLICAFR,LACCASECDNA,RTPCR引言漆酶G708LACCASE，LACG709G7171G980G12193G3822G18222G8699G2282酶G15G3324结G7512G2656G2163G14033G990与G7905物G6251G3363G15892G18252G8699G2282酶G451G2766G1095G2172物G15892G8986G19120G15025G15519G11345G4396G3324G11540G16780G3822G11468G1296G1055G3800A34A35A36。G4439G7171G7380G12628G2345的G3822G19120G8699G2282酶G15G980G14336G2559G73894G1022G19120G2419G4388G15G2010G5079于3G1022G20652度G1457G4444的G993G2528结G2524G1313G9869G15G8611G1022G19120G2419G4388G3324G1664G2282G7438G2058G1025G18129G7389G5468G18337要的作用。漆酶G14033G1664G2282G50A37G17902G178194G1022电G4388G17836G2419G6116G8712G15G5194G1000G1288G19555G11540G980G1135G18222G12879G5225物的G8699G2282A34A37A36。G4439G7380G7101G7171G1186漆G7653的G2010G8864物G1025发G10628的G15G19555G2530G1166G1216发G10628G980G1135G20652G12573G11507菌G1075G14033G2010G8864G17837G12193酶。G10628G3324G1166G1216G11705G17959G15漆酶G5203G8879G4396G3324于G6297G4388菌G451半G11705菌G2656G4388G3230菌G1025G15G1306G1866G1025G7380主要的生产G13785G7171G6297G4388菌G1025的G11345G14116菌A34A38A36，粗毛栓菌G708TRAMETESGALLICAG709G7171G980G12193G11345G14116G6297G4388菌，G3324G6117G3281G2010G5079G5203G8879，G7171G7484G7420的G980G12193主要生物G19489G16311G13785。G5062G7389研究表明，该菌产漆酶G14033力较G20652A34A39A36。漆酶G7101期的研究主要G19610G1025G3324G1319内G2163G14033的研究，G19555G115402G19世纪7G19年代前G2530聚丙烯酰胺凝胶电泳G2656G2010G4388克隆G12573生物学技术的问世G15漆酶的研究G1075揭开了新的G980页。G3324G3281外G5468G3822菌G12193的漆酶基因得到了克隆G5194G1000进行了G2010析G2656鉴定G15G980G1135漆酶基因G1075实G10628了G1866异G9316表达A34A40A36。漆酶G3324G7420质G13044G19489G16311G451微生物菌G1319形态形G6116G12573方面具G7389G18337要G2163G14033A34A40A36，普G17902微生物产漆酶的量及纯度G993足以满足大规模的工业应用。要工业G2282生产漆酶，可G17902G17819把可G20652效产漆酶的基因与载G1319G18337组，然G2530导入受G1319细胞，形G6116稳定转G2282G4388，大批量培养，使漆酶进行异G9316表达。因此必须进行漆酶基因的测序，了G16311基因组G6116及酶切G1313G9869G12573，以便DNAG18337A41A42A43A44A45A46A47A48A49A50A51组时选择G2524适的限G2058性内切酶进行G7389效切割。所以漆酶的模板CDNA提取及大量扩增G6116为首要工作。传统的模板DNA提取方法G7171采用去垢剂如SDSG12573G7481破G3363细胞组G2010，溶G16311细胞膜，使G15519G11345质变性，再用G15519G11345酶KG7481消G2282去除G15519G11345质，尤G1866G7171与DNA结G2524的G15519G11345质，再用G18222氯仿抽提，然G2530用乙醇沉淀核G18252，供PCR实验用A52A53A54。本实验主要G7171扩增G7389效表达漆酶的CDNAG10267G8585，因此利用逆转录的方法G2058备了模板CDNA，G5194对G1866进行PCR扩增，此法去除了内G2559G4388的干扰，G14033满足特殊的实验要求。CDNA完整序列的获得对基因结G7512G451G15519G11345质表达G451基因G2163G14033的研究至关G18337要。1材料与方法11主要仪器及试剂用品111主要仪器5417R型EPPENDORF离心G7438G708德G3281G709，DYY8C型G8712平电泳仪G708北京市六G980仪器厂G709，DYYIII31A型G8712平电泳槽北京市六G980仪器厂，WFH201B型G13055外G17891G4568G2465G4568仪G708G990G9035G12946科实业G7389限G1856G2508G709，干式G5670G9213器G708G7489G5042G3897G11439仪器G7389限G1856G2508G709，TP600型G7811度PCR仪G708G7097本TAKARAG1856G2508G709，HSP250生G2282培养G12677G708G2716G4584G9404市G1008G13864电G4388技术开发G7389限G1856G2508G709，GZXGFCG1031011BS电G9921G5670G9213G21735G20130干G10169G12677G708G990G9035G2350G8900实验G16786备G7389限G1856G2508G709，TU1810G13055外可G16277G2010G1821G1821度G16757G708北京G16901析G17902用仪器G7389限G17143G1231G1856G2508G709，微G8886G9821，HZQCG12366G8680G9032G6403G14645器G708G2716G4584G9404市G1008G13864电G4388技术开发G7389限G1856G2508G709。112主要试剂05DEPCG8712G708G1120乙基G9978G11911G18252G11428,DIETHYLPYROCARBONATEG709，1PBSG708G33241000MLG9793菌的去离G4388G8712G1025G2559NACL8G,KCL02G,NA2HPO4115GG2656KH2PO402G，用G11428G18252G16855PH为74，121G263，G9793菌20MING709,RNA抽提变性G909425G异G11839G8708G13985G2164入35MLCBSG13543G1926G9094G70801MOLG812LG7604G8320G18252G19060G13543G1926G9094，PH40，2SDS，1G163G5059基乙醇G709G1025，G18222氯仿G9163G2524G9094G708G18222G297氯仿G297异G6110醇25241G709，异丙醇，75乙醇，DDH2O，DH2O，琼脂糖，5G104甲醛胶G13543G1926G909401MOLG812MLMOPS，PH70G72740MMOLG812L乙G18252G19060G7275MMOLG812MLEDTA，PH80，37甲醛，G4627G13044，G2559G9352以G19201G70805G173GG812MLG709的01MOLG812L乙G18252G19145溶G9094，RNA抽提用的TAKARA试剂G11430G708G7097本TAKARAG1856G2508G709，EDTAG727琼脂糖G708G14533G3281G709，MOPSG7593G708MORPHOLINOG709PROPANESULFONICACIDG761，G2465转录试剂G11430G708G7097本TAKARAG1856G2508G709，PCR扩增试剂G11430G708G7097本TAKARAG1856G2508G709，TAEG13543G1926G9094，250BPDNALADDERMARKERG708G7097本TAKARAG1856G2508G709，G1866G4439试剂G3355为G3281产A55113主要用品G72620MLG10639G10839研G11964器，1000MLG19193形G10954，EPG12661，20G173LG451100G173LG4511000G173LG4515000G173LG7550G3848，G12239G9094G7550规G7696G72620G173L，100G173L，1000G173L,500G173L，G9793菌G10285G12626，G9793菌G9400G13452，G10639G10839G7846，G19230G4388，G980G8437性G6175G3883G451G2487G13629G451G5137G4388，G11719G14533G8616G14406G11411，量G12582G708100MLG709，G12239G9094G12661G70810MLG709，A56A57A58A59A60A61A62A63A64A65A67G9915G749112菌株及培养方式121菌株G11345G14116G6297G4388菌粗毛栓菌G13007导师孙迅于1G28G287年G27G7388G3324G4677G1008G11477G14755G8913市北G18078的G6264G3490G3576G990，G1186G15999G11745G1252的G7484G7653G990G2010离得到的。该G18338生菌株于G2528年G13475G11013G4677G1008G11477科学G19510生物研究所微生物G2010G12879G4472的G20544启明先生对G1866进行了初步鉴定，定名为TRAMETESGALLICAA68A69A70FR122培养基G726PDA培养基G726去皮土豆，200GG2164G8712煮沸20MIN,冷却至G4472G9213纱G5079G17819G9400的G9400G9094G709G727葡萄糖，20GG727琼脂粉，15GG727补G8712至1000ML。按G990述配方配G2058，G9163G2524G3355匀，取10G1022培养G11411G265620支10ML的试G12661，放入G20652压蒸汽锅G1025G9793菌G708121，30MING709结束G2530取出，待培养基冷却50G2530G2010装至10G1022培养G11411G265620支试G12661G70810MLG1025，摆斜面，冷却至G4472G9213G2058G6116固G1319培养基。浅层静置培养基半纤G13512G13044，3GG727酒G11719G18252G19145，15MGG727MG2SOA71G1037H2O，150MGG727KH2PO4，015GG727酵母粉，30MGG727100G104微量元G13044母G9094，3ML此培养基G7171G726G8616为G726G20652G11911G1314G8706培养基，适于粗毛栓菌G1025G7420聚糖酶的培养。按G990述G8616例准确称取各药品置于50ML的小G9915G7491G1025，然G2530用去离G4388G8712将G17837G1135药品溶G16311G5194定容150MLG708PH约为65G709，将定容好的培养基溶G9094G3355G2010至G1016G1022250MLG19193形G10954G1025，即G8611G1095450ML。用纱G5079及塑料G13452封好G10954G2487G2530放入G20652压蒸汽G9793菌锅G1025G9793菌G708121，30MING709，结束G2530取出G19193形G10954放至G4472G9213下冷却。冷却至35G263左右时，G3324无菌G4472内用接G12193铲G1186培养了1G19D的G12193G4388培养基G1025挖取三块米粒大的粗毛栓菌菌G1009G1319G15放入此培养基G1025，即接G12193。然G2530于26G263G5670G9213G12677进行培养，G5194于1G19DG2530G6922获菌G1009G1319。13PCR引物总CDNA的PCR扩增的引物G726OLIGODTLACF5GCTCATCGGGCCTAGTCGCA3LACR5TTACTGGTCGTCAGGCGA3以G990引物G7171G11013导师孙迅对G11507菌漆酶G2528工酶基因编码区进行G2528G9316性G8616较G2656G2010析G2530，根据编码区G1025的G1457G4444序列所G16786G16757的G12628G5194引物G708G11013TAKARAG1856G2508G2524G6116G709。14总RNA的提取与检测141总RNA的提取使用CHOMCZYNSKIG2656SACCHIA72A73A74所提出的G18252G18222抽提法G5194略作改G2172。1将培养G9094G1025的菌G1009G1319转G12239至三层纱G5079G990，G5194用G990述无菌预冷PBSG13543G1926G9094G2465复G1926洗。A76A77A78A79A80A81A82A83A84A85A862取出菌G1009G1319，置于无菌的G3822层G9400G13452G1055间，吸干G3822余的G9094G1319。3用G9793菌G10285G12626迅速将菌G1009G1319大约04G转G12239至G5062G13475DEPCG3800理G17819的20MLG10639G10839研G11964器G1025，G2164入6ML预冷的RNA抽提裂G16311变性G9094G1025研G11964，直至研G11964G9094呈G3355匀的G8986G9094状。4G2164入12ML2MOL/L的HACNAACG708PH40G709G13543G1926G9094，G9163G2524G3355匀。5将研G11964G9094转G12239至10只用DEPCG3800理G17819的15MLEPG12661G1025，向G8611只G12661G1025G2164入与样品G12573G1319积的G708约06MLG709G18222氯仿G9163G2524G9094G708G18222G297氯仿G297异G6110醇25G29724G2971G709，G9163匀，强力G6403G14645G708涡旋G70920SEC，然G2530静置冰G903210MIN，离心G70812000RPM，4G263，20MIN。6取出G990层G8712G11468至另外5只G13475DEPCG3800理G17819的15MLEPG12661G1025，向G8611只G12661G1025G2164入与样品G12573G1319积的G708约05MLG709氯仿，离心G70812000RPM，4G263，1015MING709。7取出G990层G8712G11468至另外2只G13475DEPCG3800理G17819的15MLEPG12661G1025，向G8611只G12661G1025G2164入与样品G12573G1319积的冰冷的异丙醇，G990下G2465转几G8437G708G9163匀G709，20G263冷冻1020MIN，离心G70812000RPM，4G263，15MING709，弃去G990清夜。8再向G8611G12661G1025G2010别G2164入1ML75的乙醇，离心G70812000RPM，4G263，15MING709，弃净乙醇，G4472G9213下开盖静置510MIN。9G18337复G17819程G7084G709G7088G709再抽提纯G2282G980G8437。10G2164100LDEPCG8712，G5194于20G263下G1457G4396，备用。142总RNA纯度与浓度的检测取25LG990述总RNA的提取G9094G265625ML去离G4388G8712，放入G11719G14533G8616G14406G11411G1025，G9163匀，G2010别G3324260NMG2656280NMG8886长下，用TU1810G13055外可G16277G2010G1821G1821度G16757测量G1866G1821吸G6922值，G17902G17819G16757算A260G2656A280的G8616值测G1866纯度G727G5194根据以下G1856式G16757算RNA样品浓度，总RNA的浓度G708G/LG709OD260核G18252稀释倍数401000。143总RNA的甲醛变性琼脂糖凝胶电泳A87A88A89称取04G琼脂糖于248MLG9793菌G8712G1025，G2164G9921融G2282，冷却至60G263G2530，G2164入8ML5G104甲醛胶G13543G1926G909401MOL/LMOPS3G708MORPHOLINOG709PROPANESULFONICACID,PH70G72740MMOL/L乙G18252G19060G7275MMOL/LEDTA，PH80G265672ML37甲醛，G9163匀，冷却至约50G263时，倒入G2058胶槽G1025，使凝固。取30LRNA样品与133L5G104甲醛胶电泳G13543G1926G9094，233L37甲醛G2656667L10MOL/L的G4627G13044G9163G2524，65G263，G1457G921315MIN，于冰G9032G1025速冷，G2164入15L10G104G990样G13543G1926G9094，G9163匀，立即G990样G708凝胶G5062预电泳5MING709，G33241G104甲醛胶电泳G13543G1926G9094G1025G5670压G7085V/CMG709电泳，待G9352G18222G15025染料到达凝胶G5225部约1/5G3800时，G1584G8502电泳，将凝胶G9036入G2559G9352乙G19201G70805G/MLG709的01MOL/L乙G18252G19145溶G9094G1025染G1440630MIN，G2530将G1866放入G7389MGCLA90的DHA90OG1025G9036G88853G2010G19059，G7380G2530用WFH201B型G13055外G17891G4568G2465G4568仪G7186G1699，G6305G10043。15RTPCR151逆转录及PCR利用G7097本TAKARAG1856G2508生产的G2465转录试剂G11430进行逆转录。逆转录及PCR所用试剂G11430配A91A92A93A94A95A96A97A98A99A100A101方如下G726G3324G195G80G47G40PG12661G1025G2164入G990表G1025的试剂G708此步实验样品RNA为本G4472G1457G4396的RNAG709，于干式G5670G9213器G10255G19G263G1457G9213半小时。G9213G2656G3332G9163匀G2530，G12257G12257离心，放入G7811度PCR扩增仪G1025，进行PCRG2465应，G7477G1226如下G726G284G2633G80IG81G708预变性G709G727G284G2633G19SEC，4G28G2635G27G263G708G20104G1022G7811度，G2010别G33241G4515G451G27G45112行G7093G19SEC，72G2631G80IG812G19S，G990述G18613G19G1022G5502G10627G72772G2635G80IG81G7274G263G5670G9213G1457G4396。152PCR扩增产物的琼脂糖凝胶电泳PCRG2465应结束G2530，称取075G琼脂糖于50ML05TAEG13543G1926G9094G1025，G2164G9921融G2282，冷却至约50G263时，倒入G2058胶槽G1025，使凝固。取25LPCR扩增产物与5LG990样G13543G1926G9094内G2559G9352粉G15025G708G15025G13055G14406G709G2656G1120甲G14531G19750G708G15025G14406G709G96，G9163匀，立即G990样凝胶G5062预电泳5G80IG81G64G9869G3247G17959，G8611G179591G19G173G47，DNAG80ARKERG708G8611G1022G2164样G439275LG709G2528时G990样于样品G17959G7061G17805G96，G332405TAE电泳G13543G1926G9094G1025G5670压G7085V/CMG709电泳。待染料到达凝胶G5225部约1/5G3800时，G1584G8502电泳，将凝胶G9036入G2559G9352乙G19201G70805G/MLG709的去离G4388G8712溶G9094G1025染G1440630MIN。G7380G2530用WFH201B型G13055外G17891G4568G2465G4568仪G7186G1699，G6305G10043。2结果与讨论21总RNA电泳结果G80RNA的质量G708G2010G4388的完整性G2656纯度G709G3324G19555G2530的G2465转录G2465应G1025，G7171G1927定G14033G2554获得G1852长CDNA的关键因G13044G1055G980。G11013于RRNAG3324总RNAG1025G2559量G3822G13792G12193G12879G4581，所以根据G4439G1216的G4506度G2656G2010G4388大小，G17902G17819G4506度G7811度离心G451凝胶电泳G6122离G4388G1144G6454层析G4613可以将G4439G1216G2010离。G4625G12661G80RNAG2010G4388G12193G12879G13333G3822，G1306G7171G1866G2559量G4581，G2010G4388量大小G1075G5468G993G3355G980，所以G3324电泳G3282G16901G990，G5460G5460呈微G5381的G5369G6967形G2010G5079。对新G2058备的总RNAG1025G80RNA质量的检测，G17902G5132G7171G1393G17194凝胶电泳技术，G17902G17819对RRNA质量的G2040G7041G13792确定的。G20652质量的总RNAG2058备物的电泳G3282G16901应G5415G7171，2G27SRRNA试剂G1319积G708G173G47G7091G19G104G50G81ESTEG83RTPCRG37G88G73G73ERG50G81ESTEG83G40G81G75AG81CERSOLG88TIOG81DNTPG48IG91TG88REG7081G19G80G48EACG75G709RNA102A103A104G44G81G75IBITORG7084G19G753G812G173G744G709PRIG80ESCRIG83TA105A106RTASEG73OR1STEG83TAKARAEXTAQA107A108HS5G56G812G173G47G990G9228特异性引物G7082G19G173G48G709下G9228特异性引物G7082G19G173G48G709实验样品RNA102A103A104FREEDG43A109G501G192421222273A110A111A112A113A114A115A116A98A99A100A101G26561G27SRRNA的G5114形整G21796，G1000无G14085G4626G6122G5369G6967G10628G16949G727G7389时，2G27SRRNAG5114的G1154度可达到1G27SRRNA的2倍。G3324总RNA的G2058备G17819程G1025，如果G11013于G6817作G993G5415G6122G11013于RNASE的G8757染G13792G17908G61162G27SRRNAG26561G27SRRNA部G2010G19489G16311，G18039G1052，G3324电泳G3282G16901G990，G4439G1216的G5114形G4613G1262出G10628G1017G18337G6314G4626G6122G5369G6967G10628G16949，G10990至G993G6116G5114形，G980G7098出G10628G17837G12193G5785G1929，G4613G16840明G80RNA的质量G1075G5062G13475G8821G7389G1457G16789了，即G993G14033用于RTPCR研究工作。因为G3324总RNAG1025，G80RNA应G5415G7171G1288G19555G115402G27SRRNAG26561G27SRRNA的G19489G16311G13792G19489G16311的，尤G1866G7171G11013于RNASE引G17227的G19489G16311作用G7368G7171如此A117A118A119。本G8437实验结果电泳G5194G7422出G10628理G5831的G3282G16901，G2419因G2010析可G14033主要G7171G11013于RNA样品G3324抽提G17819程G1025G3324RNA102A103A104作用下裂G16311G6122匀G8986G3800理G993G5455G5225G6122G13785G7171总RNA量G3838G4581所致。22总RNA浓度与纯度G332426G19G81G80G8886长下，1G50D值的G1821G4506度G11468G5415于G2345G19154RNA浓度的4G19G173GG18G80G47，所以本实验所提总RNA样品的浓度为G726G50DA109A120A121G104核G18252稀释的倍数G1044G19G1141G19G19G19G32G1946G104G70825G19G19G1825G709G1044G19G181G19G19G19G32G191G274GG18G47，G3324G2465转录时，4G19G47G2465转录G9094G1025的总RNA的浓度为G191G274G10425G184G19G32G19115GG18G47。测得G50DA109A120A121，G50DA109A122A121值G2010别为G1946G2656G1932G15所以G50DA109A120A121G18G50DA109A122A121的G8616值为144，G16840明总RNA纯度较G1314，G13475G2010析G2419因G5468G7389可G14033G7171因为样品G1025G9163G7389G18222G2656G15519G11345质的G13548G6937。23PCR扩增结果G11013于G1055前RNA提纯效果G993理G5831，所以G3324进行RTPCRG17819程G1025所用的RNA为本实验G4472G1457G4396的RNA，PCR扩增结果如下G726A123A124A125A126A127A128A129A130A131A132A133A134A135A136A137A138A139A140A141A142A143A144A145A140A141A146A145A144A147A140A141A146A146A144A142A140A141A146A143A144如G3282G1025可以G11487出，G3247G1022G9213度G7811度G1025，4G179595G27G263时G7477G5114G7380为明G7186，G16789明G33245G27G263时PCR扩增效果G7380好。扩增出G7481的G1016G7477CDNA长度G980G7477约为145G19BG83左右，另G980G7477约为12G19G19BG83A110A111A112A113A114A115A116A98A99A100A101左右。RNAG8616DNAG6947G5875，G7143受G19489G16311，G13792RNA酶却G8616DNA酶G2010G5079G7368G5203G8879，G8975力G7368强，G980G14336煮沸RNA酶510MIN，G186675的G8975性G1185然G8543G4396，所以G3324RNA的提取G1055前，必须对所要用的实验仪器