Yamamolekhula izindlela zocwaningo yemvelo kanye nokusetshenziswa yabo

Yamamolekhula begazi izindlela zocwaningo ukudlala indima ebalulekile kwezokwelapha zanamuhla, isayensi forensic, nezesayensi yezinto eziphilayo. Ngenxa yentuthuko kwezokuhlolwa kwezinkanyezi isifundo of DNA ne-RNA, umuntu uyakwazi ukuhlola izakhi zofuzo umzimba ukucacisa ejenti causative, ukuqaphela oyifunayo nucleic acid nenye acid, njll

izindlela Yamamolekhula-begazi. Kuyini lokhu?

Emuva e-70-80s yososayensi waba ngowokuqala wokufunda izakhi zofuzo zomuntu. Lo mcimbi kwawuqhubekisela ekuthuthukiseni ubuchwepheshe bokushintsha izakhi zofuzo kanye molecular biology. Isifundo izakhiwo of DNA ne-RNA kuye kwasho ukuthi manje kungenzeka ukusebenzisa lezi acid nucleic ngenjongo uphethwe isifo, cwaningo zofuzo.

Ukulungiselela of DNA ne-RNA

Yamamolekhula begazi izindlela zokuxilonga zidinga ukuqala impahla: Ngokuvamile lokhu nucleic acid. Kunezindlela eziningana ukuze ukhethe lezi izinto emangqamuzaneni eziphilayo. Ngamunye kunezinzuzo zako kanye nebubi, futhi kufanele kubhekwe lapho ekhetha indlela ukuzihlalela acid nucleic ngefomu okumsulwa.

1. Ukulungiselela ye-DNA yi-Marmur. Indlela liqukethe ekwelapheni ingxube utshwala izinto, ehlele okuholela DNA okumsulwa. Yini embi ngalo le ndlela kuyinto ukusetshenziswa izinto ezonakalisayo, phenol futhi chloroform.

2. Kwemihlambi DNA phezu boom. Ingqikithi oyinhloko elisetshenziswe lapha - kuyinto guanidine thiocyanate (GuSCN). Ukhuthaza sebethule deoxyribonucleic acid ku substrates ayisipesheli, okuyindawo kungenziwa kamuva eqoqwe esebenzisa sesikhashana ekhethekile. Nokho GuSCN - kuyinto inhibitor we-TCP, ngisho ingxenye encane ukuthi ihlala ezimpapheni e-DNA bangathonya Yiqiniso ukuthi bayavuma polymerase chain, okuyinto ebaluleke lapho esebenza acid nucleic.

3. Inzika amanyela. Indlela iyahluka kwenye abathandekayo sangaphambilini ukuthi amangqamuzana bona abawona imali dehoksiribonukleinovoy acid amanyela. Ukuze wenze lokhu, sebenzisa kwabebhange ion. Ububi ukuthi akubona bonke izinto kungaba axazulule.

4. IMisa Ukuhlolwa. Le ndlela isetshenziswa ezimweni lapho awudingi ukuba ulwazi olungenaphutha ngemithi isakhiwo se-molecule ye-DNA, kanye nesidingo ukuthola izibalo ezithile. Isizathu wukuthi nucleic acid isakhiwo kungalimala lapho usingatha insipho, ikakhulukazi alkalies.

Ukwahlukanisa izindlela zocwaningo

Zonke izindlela yamangqamuzana-begazi ucwaningo zihlukaniswe amaqembu ezinkulu ezintathu:

1. okhulisa (usebenzisa sebuningini enzyme). Lokhu kuhlanganisa i-PCR - polymerase chain reaction, edlala indima ebalulekile eziningi izindlela zokuxilonga.

2. Neamplifikatsionnye. Leli qembu izindlela lihlotshaniswa ngqo umsebenzi nucleic acid Izingxube. Izibonelo 3 izinhlobo usula, in situ hybridization, njll

3. Izindlela esekelwe ukuqashelwa yengeza izitshengiselo ezivela kwamanye engqamuzaneni uphenyo, okuyinto sibopha a DNA noma RNA uphenyo oluthile. Isibonelo - hybridization uhlelo isixazululo Hybride Capture System (hc2).

Enzyme ukuthi ingasetshenziswa yamangqamuzana izindlela zocwaningo yemvelo

amasu amaningi yamangqamuzana zokuxilonga ezihilela ukusetshenziswa ezibanzi enzyme. Ngezansi asetshenziswa avame:

1. Umkhawulo enzyme - "usike" molecule ye-DNA kuzingxenye kunesidingo.

2. DNA polymerase - synthesizes a kabili balahleke deoxyribonucleic acid-molecule.

3. transcriptase Ukuhlanekezela (ukuhlanekezela transcriptase) - esetshenziswa synthesize DNA kusukela isifanekiso RNA.

4. DNA ligase - unesibopho kumiswa phosphodiester isibopho phakathi ama-nucleotide.

5. exonuclease - kususa nucleotide kusukela izingxenye ekupheleni deoxyribonucleic acid-molecule.

PCR - indlela eyisisekelo DNA okhulisa

Polymerase chain reaction (i-PCR) is kabanzi molecular biology yesimanje. Le ndlela, lapho DNA olulodwa ingatholakala isibalo esikhulu sezithunjwa amakhophi (i-molecule yokukhulisa izwi).

Imisebenzi esemqoka PCR:

- izifo;

- cloning izingxenye DNA, Gene.

Ukuze afeze ukusabela polymerase chain kudinga izinto ezilandelayo: kokuqala molecule ye-DNA, i-polymerase DNA thermostable (Taq noma Pfu), phosphate deoxyribonucleotide (imithombo nitrogen lezisekelo), le kwenziwa (2 primer 1 DNA) kanye uqobo uhlelo sesikhashana okungase ukuqhuba zonke ukusabela.

PCR siqukethe izinyathelo ezintathu: denaturation, annealing primer kanye ukunwebeka.

1. denaturation. At lokushisa 94-95 degrees Celsius proshodit baphule izibopho hydrogen phakathi namaketanga amabili DNA, futhi ngenxa yalokho sithola molecule ezimbili owodwa balahleke.

2. kwenziwa annealed. At lokushisa 50-60 degrees centigrade kwenzeka ukubusa kwenziwa enisemikhawulweni nucleic acid molecule elilodwa-balahleke ngokuvumelana uhlobo complementarity.

3. ukunwebeka. At lokushisa 72 degrees is indodakazi kwemiqondo double balahleke deoxyribonucleic acid molecule.

• Ukutshengisa ukulangazelela nokuthatheka DNA

Yamamolekhula izindlela zocwaningo begazi ngokuvamile zidinga ulwazi ukulandelana nucleotide e-molecule deoxyribonucleic acid. Ukuze sithole isimiso sezakhi zofuzo zilandelaniswe khona. ukuhlola yamangqamuzana Ngokuzayo kuyoba ngokohlelo lweSitatimende lwati lolutfolakele ekutholweni ukulandelana womuntu.

Lezi zinhlobo ezilandelayo • Ukutshengisa ukulangazelela nokuthatheka:

• • Ukutshengisa ukulangazelela nokuthatheka yi Maxam-Gilbert;
• Sanger ngekulandzelanisa;
• pyrosequencing;
• nanoporovoe kulandzelanisa.