Robustart Taq DNA Polymerase

Robustart Taq DNA Polymerase bụ mmalite DNA polymerase na-ekpo ọkụ.Ngwaahịa a nwere ike ọ bụghị naanị ka mma igbochi mmeghachi omume na-abụghị nke akọwapụtara nke ọma site na mkpochapụ na-enweghị isi nke primers ma ọ bụ nchịkọta primer na usoro nkwadebe na nkwalite usoro PCR.Ya mere, ọ nwere nkọwa dị mma ma dị irè karị maka mmụba nke ndebiri ntinye uche dị ala, ọ dịkwa mma maka mmeghachi omume mmụba PCR multiplexed.Ọzọkwa, ngwaahịa a nwere ezigbo ngwa ngwa, yana nsonaazụ mmụba kwụsiri ike nwere ike nweta n'ụdị mmeghachi omume PCR dị iche iche.

Ngwa

1.5 U/μL Robustart Taq DNA polymerase

2.10 × PCR Buffer II (Mg²+ n'efu) (nhọrọ)

3.25 mmMgCl₂(nhọrọ)

* 10 × PCR Buffer II (Mg²+ n'efu) enweghị dNTP na Mg²+, biko tinye dNTP na MgCl₂mgbe ị na-akwadebe usoro mmeghachi omume.

Ngwa akwadoro

1.Ngwa ngwa nkwalite.

2.Otutu nkwalite.

3.Mmụba ọbara, swabs na ihe nlele ndị ọzọ ozugbo.

4.Nchọpụta ọrịa iku ume.

Ọnọdụ Nchekwa

-20ºC maka nchekwa ogologo oge, ekwesịrị ịgwakọta nke ọma tupu ejiri ya, zere ịza oyi mgbe niile.

*Ọ bụrụ na mmiri ozuzo na-eme mgbe refrigeration gasịrị, ọ bụ ihe nkịtị;a na-atụ aro ka ọ dakọtara na ọnụ ụlọ okpomọkụ tupu ịgwakọta na iji.

Nkọwa nkeji

A kọwapụtara otu nkeji na-arụ ọrụ (U) dị ka ọnụọgụ nke enzyme nke na-etinye 10 nmol nke deoxyribonucleotide n'ime ihe anaghị emerụ acid na 74 Celsius C maka nkeji 30 n'iji DNA sperm salmon arụ ọrụ dị ka template/primer.

Njikwa ogo

1.SDS-PAGE electrophoretic dị ọcha karịa 98%.

2.Mmetụta mmụba, njikwa batch-to-batch, nkwụsi ike.

3.Ọ dịghị ọrụ nuclease exogenous, ọ dịghị exogenous endonuclease ma ọ bụ exonuclease mmetọ

Ntuziaka

Ntọala mmeghachi omume

ndetu:

1) a.Ihe nchekwa ahụ enweghị dNTP na Mg²+, biko tinye dNTP na MgCl₂mgbe ị na-akwadebe usoro mmeghachi omume.

2) b.Ọ bụrụ na ejiri ya maka qPCR/qRT-PCR, ekwesịrị ịgbakwunye nyocha fluorescent na sistemụ mmeghachi omume.Na-emekarị, njedebe ikpeazụ nke 0.2 μM ga-enye nsonaazụ dị mma;ọ bụrụ na mmeghachi omume mmeghachi omume adịghị mma, enwere ike ịhazi ntinye nke primer na oke nke 0.2-1 μM.A na-ebukarị mkpokọta nyocha na oke 0.1-0.3 μM.Enwere ike ịme nnwale gradient ntinye uche iji chọta ngwakọta kacha mma nke primer na nyocha.

Usoro ịgba ígwè okpomọkụ

Ihe ndetu

1.Ọnụego mmụba nke DNA polymerase ngwa ngwa ekwesịghị ịbụ ihe na-erughị 1 kb/10 s.Ọnụego ịrị elu na ọdịda okpomọkụ, ọnọdụ njikwa okpomọkụ na nrụpụta okpomọkụ nke ngwa PCR dị iche iche dịgasị iche iche dị iche iche, ya mere a na-atụ aro ka ebuli ọnọdụ mmeghachi omume kachasị mma maka ngwa ngwa PCR ngwa ngwa.

2.Usoro a na-agbanwe nke ukwuu, yana nkọwa dị elu na nghọta.

3.Kwesịrị ekwesị maka ojiji dị ka reagents nchọpụta PCR nwere uche dị elu, enwere ike iji ya na nzaghachi mmụba PCR multiplex.

4.5′→3′ ọrụ polymerase, 5′→3′ ọrụ exonuclease;ọ dịghị 3′→5′ exonuclease ọrụ;enweghị ọrụ nyochagharị.

5.Kwesịrị ekwesị maka ule qualitative na quantitative nke PCR na RT-PCR.

6.Ọgwụgwụ 3′ nke ngwaahịa PCR bụ A, nke enwere ike itinye ya ozugbo na vector T.

7.A na-atụ aro usoro nzọụkwụ atọ maka primers nwere obere okpomọkụ na-ekpo ọkụ ma ọ bụ maka mmụba nke iberibe ogologo karịa 200 bp.