8һøԶAla-GlyͶLeu-GlyKmֱΪ10-4͡10-2һøʵøǾƼKiֱֵΪ10-2͡10-4һǿƼ
𣺿ɸϷ̼ƼʱøٷӦʷжϣ KmֵСAla-GlyʵKiֵСǸǿƼ
91/v1/[S]˫ͼõֱбΪ10-3min1/vϵĽؾΪ10-2nmol-1ml minVmaxKm
бS=(1/ Vmax)(1/ Km)= Km/ Vmax=10-3min,Ϊ
1/Vmax=10-2nmol-1ml minԣ Vmax50 nmol ml-1 min-1 Km102 nmolml-1
DNAԵԡ
DNAıָDNA˫ĶۺѣɵṹĹ̡DNAϡҺȵ80-100ʱ˫ṹ壬˴˷ֿγšDNAԺһϵҲ֮仯ճȽ͡(260 nm)ֵߵȡ ϰ һʽ
:ϵӢдDNAƹнڸսĵϷֹ½ϻDNAø⡣ 2. Exon:ӣָѻЯŴϢDNAƬΡ
3. ṹ:ɵʵĶṹ۵γɵĽڵʶṹṹ֮һֵʽṹ
4. Ͳ:ָõʷӶеʶͽķ봿
5. ṹ:ڳṹṹ֮һֵʿռṹɵʶṹһ۵γɡ
6. :DNA˫ЯŴϢ
7. нṹ:RNAͷӿͨγɾֲ˫ṹ˫ֺͷ˫״ɵĽṹƷУΪнṹ
8. :ҺӴκֵĵʳ 9. ɫЧӦ:˫ɵ260nmӵ
10.ʽ:˳ʽԪԽϵĵӱΪʽӡ 10. ʴ
1.ӽӡм֣ǷֱмҪ˵
𣺢SouthernӡӾDNAƬתάĤϺ뾭ǵDNA̽ӽ
NorthernӡRNAӾԺתάĤٽӽ
WesternӡݿͿԭԽϵԭúӽƷеʷ 2.øŨNaClҺܽԼӳķԴϸȽзDNA˵ʵ鲽ԭ
ͨϸеĺʶ뵰ʽһγɺ˵סDNAڸŨȻҺڵŨȵȻҺRNA෴һ㽫˵뵽1ĦŨȵȻҺпԽDNARNA뿪ñȡȥסˮ͵ıDNA䶳ġʹѸٱԣ˾DNAϲˮСαӴͿԵõȻ״̬DNA
3. ΪʲôDNARNAԲöǷкⶨԸԲⶨľ巽Ϊ˵
𣺢ٺǺIJⶨRNAеĺǺŨŨˮɿȩȩ·ӷӦɫӦ660nmգRNAŨȳȡ
ǺIJⶨDNAеǺŨˮɦ-ǻ--ͪȩ
ӦɫӦ595nmգDNAŨȳȡ
4. ϸµľƾУʹ÷Աǵ̼Դʾԭʵ顣ʼ
14
ǷӵʲôλϱC ʹҴǻ̼ǻ̼ԭӣλϣʹþƾҪʽ˵
5. øܹмࣿǸԴַӦ
߱ʾҪ˵Ũȡ¶ԼpHֵøٷӦٶȵӰ졣 𣺢øķԼͬøķӦ£
ԭøࣺԭӦ תøࣺܻŵתƣ ˮøࣺˮķӦ
Ѻøࣺˮ̼ȥ룻 칹øࣺ͵칹ã ϳøࣺATPijɼӦ ƲͬضøٷӦٶȵӰ£
6. ﵰʺϳԭЩҪ Ҫڣ
ٲ뷭ʼeIF϶࣬eIF2eIF3eIF4 ϸĺϴ80s
γʼİǼǼ γʼĻƲͬ
ϸmRNAΪ˳ӣԭϸmRNAΪ˳ӣ
ԭϸӳEF-TUEF-GӦϸӳΪEF1EF2
7.һδ֪ĵİΪ Asp1, Ser1, Gly1, Ala1,Met1, Phe1Lys2һϵз£
DNFB֮ӦˮõDNP-Ala
ھ鵰øӲзһģΪAsp1, Gly1, Lys1, Met1, ĵDNFBӦΪDNP-Gly
۸ðľȵøɵõɷֱΪLys1Ala1Ser1ԼPhe1Lys1Gly1ļһġ˶ľCNBr춬ᡣƵðС
𣺸ݽ٣ΪAla-Asp1, Ser1, Gly1, Met1, Phe1Lys2
ݽڣΪAla-Ser,Lys-Phe- Gly C(Asp, , Met,Lys ݽۣΪAla-Ser-Lys-Phe- Gly CLys-Met CAsp
8.ʲôͬøøΪ˵ɡΪʲôõӾøз룿˵øڼеӦá6֣
𣺴ַͬӦIJͬøΪͬøøΪ 9. ԭmRNAĽṹصʲô ԭϸmRNAṹص㣺
һΪ˳ӽṹ ת¼뷭ϵģ
mRNAӰȵͷǷ˳֮ڲУ ϸmRNAṹص㣺
ϸmRNA5@ĩñӽṹ3@ĩpolyAβͣ ϸmRNAһΪ˳ӣ
ϸmRNAת¼뷭Ƿֿеģ
10. Ƚϵۡԭάڽṹɷͬǵ ߽ΪǷͨռӶɵĶӡ
۷ֱۺֱ֧ۡǷͨ-14ռɵֱṹ֧ÿֱӼǦ-16ռˮвܽ⣬ڼȵ¿ˮֶͳɺ״ֱɫ֧Ϻɫ
ԭֳƶۣͬҲɦ-D-ǹɵͬ۶ǣṹ֧ƣҲǴЦ-16֧Ħ-14Ƕ֧֧۶ࡣԭɺɫ
άɦ-D-ͨ-14ռɵͬ۶ǡάزˮϡϡάÿDzлǻγ 11. ѪǵԴȥ·άѪˮƽĺ㶨
ѪҪԴڶʳգԭķֽԼʾǣѪǵҪȥ·dzֽͷϳԭԼںϳ֬ʡڵѪˮƽҪȵأȸѪغءȵͨýѪǣٽת˽ϸԭϳɣӿǵã֬ԱȡȸѪͨ·ʽѪǣԭϳɣٽԭֽ⣬ǽͽ⣬ٽ֬ԱҲǿѪǵļأҪͨԭֽѭʵ֡ 12.ʲôǵʵı˵ʱĹϵ
𣺵ʷܵijЩصӰ죬ռ乹ƻʧȥԭеԵʱԡʷӵܽ½ҺʳʹʳķлܼȡijЩǿҵķȣлᣬؽεʹʳĻĵʻԡȽº͵ķʱĵʲԡԵĵһЩʵϡҺᡢȵķԺ磬ţУеҵױˣûг
13. lacӵĻṹģͨCAPڻģ
𣺢 DzӽṹͼʾṹɣDZ-øĻZͨøĻYøĻa
λڽṹǰǵPͲOɡλڵǰI⣬λһζǷֽлCAPĽ
ṹصķʽ£
ʱIϣӶסRNAۺøǰƵͨ·ṹת¼
ڵǴʱΪյϣʹ乹ϣ˽ṹת¼ǵø
۵ǺǶʱϸͨȻǡʱDzӱϲʱDzӵıǿͨCAPʵֵġ
ܵcAMPCAPϺ߹仯DNAǿDz CAPĽλ cAMP-CAPϵ DNAϵ CAPλٽ RNAۺøϵϣת¼Žϸϸ cAMPĺ cAMP-CAP٣ӰDzת¼꣬cAMPʱDzӵת¼CAPһڵףҪcAMP롣 ϰߴ һʽ
2+
1. ˫巴Ӧ:ָڼҺCuϺɫķӦǵʷļķӦļԽ෴ӦɫԽ
2. ˳ʽԪ:ָλDNAϵضӺǿӵԪͨǿԿƻת¼ 3. :DNA˫ЯŴϢ
4.ǿ:ָϸзֵһЩǿӻԵĺС
5.Ľṹ:DNAijЩƬУЩijһ180תԳƵУ˳ͬ
СǻĽṹ
6.ѧ:ָоͻɣṹȶԼ书ܵĿѧ 7.:ҺУŨȵܴٽܽܡ 8.ˮ:ָǼԻΪܿˮ 9. Intron:λڶѻеIJЯŴϢDNAƬΡ
10.ɫЧӦ:ɵ˫260nm½ 11. ʴ
Ŵ˳ɶİ˳֤ȷĹؼʲô 𣺹ؼ㣺
ٰtRNAϳøȷtRNAϵԣ ijְֻضtRNAϣ ͨ뷴ԣ ضİtRNAֻضmRNAϵԽϣ ͨ㣬֤ضضӵĶӦϵҲͱ֤˷ȷԡ DNSEdmanҪʲô棿֮Ҫʲô
DNFBDNSEdmanڶN-ĩ˰ķⶨ֮⣬EdmanڶеķDNSԭDNFBԭͬȸDNFBҷӦȡͿֱӽм
EdmanDNFBDNSȣÿⶨһN-ĩ˰µĶȻɹⶨһᣬ˸÷ڲⶨС 3. еҪҪЩ 𣺵еҪҪУ ã ڴлã ߱ã ת˺ʹң ˶֧ã ֻã ߽ܺʹϢãĤĹܣ ʶܣ
4. ϸµľƾУʹ÷Աǵ̼Դʾԭʵ顣ʼ
14
ǷӵʲôλϱC ʹҴļ̼λϣʹþƾҪʽ˵
16 ٶһ·ֵIJȡ˺ᣬϤķȷ DNARNA
ǵ˫ṹд÷ʵ鲽˵
𣺢ٿͨǷȷƷŨᷴӦм·Լɫ˵ƷRNADNAڿͨɫЧӦж˫ǵȲⶨƷA260 ȻͨʹԣٲⶨA260˵ú˫˵ǵ 6.øͻѧ֮Ĺͬʲôø뻯ѧȵIJ֮ͬʲô
𣺹ͬ㣺Чʸߣڲı䷴Ӧƽ̷ֻӦƽʱ䣻۽ͷӦܣܷӦǰøʹı䡣
ͬ㣺پøǵʣøܻصӰ죻ͨøĴЧʸߣøרһԸǿøĻԿɱڣһЩĻѧӦֻͨø 7.øøЧʵĻʲô
DNA˫ṹȶʲô 𣺢øøЧʵĻǣ
2. ġ͡ЧӦ 3. ã 4. ۴ã 5. ã
DNA˫ṹȶǣ