Te Whakatau Rapanga mō te Kaitātari PCR: Ngā Pātai Auau me ngā Otinga

He taputapu nui ngā tātaritanga tauhohenga mekameka polymerase (PCR) i roto i te koiora ngota, e āhei ai ngā kairangahau ki te whakanui i te DNA mō ngā tono mai i te rangahau ira ki te tātaritanga haumanu. Heoi, pērā i ngā taputapu uaua katoa, ka tūpono pea he raruraru ki tētahi tātaritanga PCR e pā ana ki tana mahi. Ka aro atu tēnei tuhinga ki ētahi pātai noa mōKaitātari PCRte rapurongoā, me te whakarato i ngā otinga whai hua mō ngā raruraru noa.

1. He aha te take kāore taku tauhohenga PCR e whakanui ake ana?

Ko tētahi o ngā raruraru e pā ana ki ngā kaiwhakamahi ko te kore e taea e te tauhohenga PCR te whakanui ake i te DNA ūnga. Ka taea te kī he maha ngā āhuatanga e pā ana ki tēnei:

Hoahoa paraime hē: Kia tino motuhake ō paraime mō te raupapa ūnga, ā, kia tino pai te pāmahana rewa (Tm). Whakamahia ngā taputapu pūmanawa mō te hoahoa paraime hei karo i te herenga kore-motuhake.

Kāore i te Rawaka te DNA Tauira: Manatokohia kei te whakamahia e koe he nui te DNA tauira. Ki te iti rawa ka ngoikore, ka kore rānei e whakanuia.

Ngā aukati i roto i te tauira: Ka taea e ngā poke i roto i te tauira te aukati i te tauhohenga PCR. Whakaarohia te pure i tō DNA, te whakamahi rānei i tētahi atu tikanga tango.

Otinga: Tirohia te hoahoa o tō paninga, whakanuia te kukū o te tauira, ā, kia tino mohio kāore he aukati i roto i tō tauira.

2. He aha i hē ai te rahi o taku hua PCR?

Mena kāore te rahi o tō hua PCR i te rite ki te mea e tumanakohia ana, tera pea he raruraru kei ngā āhuatanga tauhohenga, kei ngā kai rānei i whakamahia.

Whakareitanga kore-motuhake: Ka puta pea tēnei mēnā ka piri te paraime ki tētahi wāhi kāore i te whakaarohia. Tirohia te motuhaketanga o ngā paraime mā te whakamahi i tētahi taputapu pēnei i te BLAST.

Te Pāmahana Whakamahana Hē: Ki te iti rawa te pāmahana whakamahana, ka puta pea he herenga kore-motuhake. Te arotau i te pāmahana whakamahana mā te PCR gradient.

Otinga: Whakaūtia te motuhaketanga o te paraime me te arotau i te pāmahana whakamahana hei whakapai ake i te tika o ngā hua PCR.

3. Ka whakaatuhia e taku tātaritanga PCR tētahi karere hapa. Me aha ahau?

He mea whakamataku ngā karere hapa i runga i te tātaritanga PCR, engari he maha ngā wā ka taea e rātou te whakarato tohu ki ngā raruraru pea.

Ngā Take Whakatikatika: Me whakarite kei te tika te whakatikatika o te tātaritanga PCR. He mea nui te tiaki me te tirotiro whakatikatika auau kia tika ai ngā hua.

Rōpū Pūmanawa: I ētahi wā, ka raru pea ngā hapa pūmanawa. Tīmata anō i tō rorohiko, tirohia mēnā he whakahōunga pūmanawa.

OTINGA: Tirohia te pukapuka kaiwhakamahi mō te waehere hapa motuhake, ā, whai i ngā mahi rapurongoā kua tūtohutia. Mā te tiaki auau ka ārai i ngā raruraru maha.

4. He aha i kore ai e ōrite ngā hua o taku tauhohenga PCR?

Ka raru pea ngā hua PCR kāore i te ōrite mō ētahi take:

Te Kounga o te Tauhohenga: Me whakarite kia hou, kia kounga teitei hoki ngā tauhohenga katoa, tae atu ki ngā whākōkī, ngā ārai, me ngā dNTP. Tērā pea ka rerekē te kounga o ngā tauhohenga kua pau te wā, kua poke rānei.

Te Whakatikatika i te Pūrere Whakamahana: Tērā pea ka pāngia te tukanga PCR e ngā tautuhinga pāmahana rerekē. Tirohia tonutia te whakatikatika o te pūrere whakamahana.

Otinga: Whakamahia ngā matū kounga teitei, ā, whakatikatikahia tō mīhini huringa wera i ia wā, i ia wā, kia rite tonu ai ngā hua.

5. Me pēhea te whakapai ake i te whai huatanga o te tauhohenga PCR?

Mā te whakapai ake i te whai huatanga o ngā tauhohenga PCR ka hua ake he hua teitei ake me ngā hua pono ake.

Arotauhia ngā āhuatanga tauhohenga: Whakamātauria mā te whakamahi i ngā kukū rerekē o ngā paraime, te tauira DNA me te MgCl2. Tērā pea ka hiahiatia he āhuatanga ahurei mō ia tauhohenga PCR kia pai ai te mahi.

Whakamahia ngā whākōkī pono-teitei: Mena he mea nui te tika, whakaarohia te whakamahi i te DNA polymerase pono-teitei hei whakaiti i ngā hapa i te wā whakanui.

Otinga: Whakahaerehia he whakamātautau arotau hei kimi i ngā āhuatanga pai rawa atu mō tō tatūnga PCR.

Hei whakarāpopototanga

Te rapurongoā i tētahiKaitātari PCRHe mahi uaua pea, engari mā te mārama ki ngā raruraru noa me ā rātou otinga ka tino whakarei ake i tō wheako PCR. Mā te whakaoti i ēnei raruraru noa, ka taea e ngā kairangahau te whakapai ake i ngā hua PCR me te whakarite kia pono ngā hua i roto i ngā tono koiora ngota. Ko te tiaki auau, te whiriwhiri tupato i ngā matū, me te arotau i ngā āhuatanga tauhohenga ngā mea nui ki te tātaritanga PCR angitu.