Kutulutsa kwa Nucleic Acid ndi Magnetic Bead Method

Mawu Oyamba

Kodi Nucleic Acid Extraction ndi chiyani?

M'mawu osavuta kwambiri, nucleic acid m'zigawo ndikuchotsa RNA ndi / kapena DNA kuchokera ku chitsanzo ndi zowonjezera zonse zomwe sizofunikira.Njira yochotsera imalekanitsa ma nucleic acid kuchokera pachitsanzo ndipo amawapereka ngati mawonekedwe osakanikirana, opanda zosokoneza komanso zowononga zomwe zingakhudze ntchito zilizonse zapansi.

Kugwiritsa Ntchito Nucleic Acid M'zigawo

Ma nucleic acid oyeretsedwa amagwiritsidwa ntchito muzochulukira zamitundu yosiyanasiyana, kuyambira m'mafakitale osiyanasiyana.Zaumoyo mwina ndi malo omwe amagwiritsidwa ntchito kwambiri, okhala ndi RNA yoyeretsedwa ndi DNA yofunikira pazifukwa zosiyanasiyana zoyesa.

Kugwiritsa ntchito kwa nucleic acid m'zipatala kumaphatikizapo:

- Kukulitsa kwa PCR ndi qPCR

- Next Generation Sequencing (NGS)

- Amplification-based SNP Genotyping

- Array-based Genotyping

- Kuletsa kwa enzyme Digestion

- Kusanthula pogwiritsa ntchito ma Enzymes osintha (mwachitsanzo Ligation ndi Cloning)

Palinso magawo ena opitilira chisamaliro chaumoyo komwe kuchotsedwa kwa ma nucleic acid kumagwiritsidwa ntchito, kuphatikiza koma osangokhala pakuyesa kwa abambo, ma forensics ndi genomics.

Mbiri Yachidule ya Kutulutsa kwa Nucleic Acid

Kutulutsa kwa DNAkudayamba kalekale, ndipo kudzipatula koyamba kodziwika kudachitika ndi dokotala waku Switzerland wotchedwa Friedrich Miescher mu 1869. Miescher anali kuyembekezera kuthetsa mfundo zazikuluzikulu za moyo pozindikira kapangidwe kake ka maselo.Atalephera ndi ma lymphocyte, adatha kupeza DNA yochuluka kuchokera ku leukocyte yomwe imapezeka mu mafinya pamabandeji otayidwa.Anachita izi powonjezera asidi ndiyeno alkali mu selo kuti achoke mu cytoplasm ya selo, kenako anapanga ndondomeko yolekanitsa DNA ndi mapuloteni ena.

Kutsatira kafukufuku waposachedwa wa Miescher, asayansi ena ambiri apita patsogolo ndikupanga njira zodzipatula ndikuyeretsa DNA.Edwin Joseph Cohn, wasayansi wamapuloteni adapanga njira zambiri zoyeretsera mapuloteni pa WW2.Iye anali ndi udindo wolekanitsa gawo la seramu la albumin la plasma, lomwe ndi lofunika kusunga kuthamanga kwa osmotic m'mitsempha ya magazi.Zimenezi zinali zofunika kwambiri kuti asilikali apitirizebe kukhala ndi moyo.

Mu 1953 Francis Crick, pamodzi ndi Rosalind Franklin ndi James Watson, anadziŵa mmene DNA inapangidwira, kusonyeza kuti inali ndi timizere tiŵiri tautali ta nucleic acid nucleotides.Kutulukira kumeneku kunatsegula njira kwa Meselson ndi Stahl, omwe adatha kupanga ndondomeko ya density gradient centrifugation protocol kuti alekanitse DNA kuchokera ku mabakiteriya a E. Coli pamene adawonetsa kubwereza kwa DNA kokhazikika mu 1958.

Njira Zopangira Nucleic Acid

Kodi magawo 4 a DNA m'zigawo ndi ziti?
Njira zonse zochotsera zimachokera ku masitepe ofanana.

Kusokoneza Maselo.Gawoli, lomwe limadziwikanso kuti cell lysis, limaphatikizapo kugwetsa khoma la cell ndi/kapena nembanemba ya cell, kuti atulutse madzi am'kati mwa cell omwe ali ndi nucleic acid.

Kuchotsa Zinyalala Zosafuna.Izi zimaphatikizapo ma membrane lipids, mapuloteni ndi ma nucleic acid ena osafunikira omwe amatha kusokoneza ntchito zotsika.

Kudzipatula.Pali njira zingapo zolekanitsira ma nucleic acids osangalatsa kuchokera ku lysate yoyeretsedwa yomwe mudapanga, yomwe imagwera pakati pa magulu awiri akulu: yankho lokhazikika kapena lolimba (onani gawo lotsatira).

Kukhazikika.Pambuyo pa nucleic acids atasiyanitsidwa ndi zonyansa zina zonse ndi zosakaniza, zimaperekedwa mwapamwamba kwambiri.

Mitundu Iwiri Yochotsa
Pali mitundu iwiri ya nucleic acid m'zigawo - njira zothetsera ndi njira zolimba.Njira yothetsera vutoli imadziwikanso kuti njira yochotsera mankhwala, chifukwa imaphatikizapo kugwiritsa ntchito mankhwala kuti awononge selo ndikupeza nucleic material.Izi zitha kukhala kugwiritsa ntchito mankhwala ophatikizika monga phenol ndi chloroform, kapena mankhwala ocheperako omwe amalimbikitsidwa kwambiri monga Proteinase K kapena gelisi ya silica.

Zitsanzo za njira zosiyanasiyana zochotsera mankhwala kuti muwononge selo ndi:

- Kuphulika kwa Osmotic kwa nembanemba

- Enzymatic chimbudzi cha khoma la cell

- Kusungunuka kwa membrane

- Ndi zotsukira

- Ndi mankhwala a alkali

Njira zolimbitsa thupi, zomwe zimadziwikanso kuti njira zamakina, zimaphatikizapo kugwiritsa ntchito momwe DNA imalumikizirana ndi gawo lapansi lolimba.Posankha mkanda kapena molekyulu yomwe DNA idzamangirizepo koma wosanthula sangatero, ndizotheka kusiyanitsa ziwirizo.Zitsanzo za njira zopangira magawo olimba kuphatikiza kugwiritsa ntchito mikanda ya silika ndi maginito.

Kutulutsa Mkanda Wamaginito Kufotokozera

Njira Yopangira Mkanda Wamaginito
Kuthekera kwa kutulutsa pogwiritsa ntchito mikanda ya maginito kudadziwika koyamba mu patent yaku US yomwe Trevor Hawkins adapereka, ku bungwe lofufuza la Whitehead Institute.Patent iyi idavomereza kuti ndizotheka kutulutsa ma genetic powamanga ku chonyamulira cholimba, chomwe chingakhale mkanda wa maginito.Mfundo ndi yakuti mumagwiritsa ntchito kwambiri functionalised maginito mkanda pa zimene chibadwa adzamanga pa, amene kenako analekanitsidwa ndi supernatant ndi ntchito maginito mphamvu kunja kwa chotengera akugwira chitsanzo.

Chifukwa Chiyani Muzigwiritsa Ntchito Magnetic Bead Extraction?
Ukadaulo wochotsa mikanda wa maginito ukuchulukirachulukira, chifukwa cha kuthekera komwe kumakhala nawo pakuchotsa mwachangu komanso moyenera.Posachedwapa pakhala pali chitukuko cha mikanda ya maginito yogwira ntchito kwambiri yokhala ndi makina oyenera obisala, omwe apangitsa kuti ma nucleic acid achuluke komanso njira yogwirira ntchito yomwe imakhala yopepuka komanso yotsika mtengo.Komanso, njira zochotsera mikanda ya maginito siziphatikiza masitepe apakati omwe angayambitse mphamvu zometa ubweya zomwe zimaswa zidutswa zazitali za DNA.Izi zikutanthauza kuti zingwe zazitali za DNA zimakhalabe, zomwe ndizofunikira pakuyesa ma genomics.