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PCR standar terdiri dari DNA target, satu set primer oligonukleotida sintetis yang mengapit sekuens DNA target, DNA polimerase termostabil (biasanya Taq polimerase), dan nukleotida. Dengan menggunakan siklus termal, ada tiga tahap selama setiap siklus amplifikasi, termasuk mendenaturasi DNA untai ganda (dsDNA) menjadi DNA untai tunggal yang terpisah, annealing primer ke sekuens DNA target, dan ekstensi, di mana DNA polimerase memperpanjang DNA dari primer, menciptakan dsDNA baru dengan satu untai lama dan satu untai baru. 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[Belanja Produk](https://www.sigmaaldrich.com/ID/id/products/molecular-biology-and-functional-genomics/microbiome) * * * Ikhtisar Artikel & Protokol Terkait Dukungan ## Reverse Transcriptase PCR (RT-PCR) RT-PCR, atau reverse transcriptase PCR, adalah variasi dari teknik PCR standar yang melibatkan amplifikasi mRNA spesifik yang diperoleh dari sampel yang sangat kecil. Ini menghilangkan kebutuhan akan proses pemurnian mRNA yang membosankan yang diperlukan untuk teknik kloning konvensional. Dengan RT-PCR, transkriptase balik dan sampel RNA digunakan sebagai tambahan untuk reagen PCR standar. Campuran reaksi dipanaskan hingga 37˚C, yang memungkinkan produksi salinan cDNA komplementer dari sampel RNA dengan transkriptase balik. cDNA ini kemudian diannealing ke salah satu primer yang mengarah ke sintesis untai pertama. PCR standar mengikuti dari sini di mana dsDNA pada akhirnya dihasilkan. RT-PCR sering dikombinasikan dengan real time PCR (qPCR), yang banyak digunakan untuk kuantifikasi tingkat transkrip dalam sel dan jaringan. PCR Mulai Panas PCR Mulai Panas adalah teknologi yang menghambat Taq polimerase mulai panas atau penggabungan dNTP yang dimodifikasi selama pengaturan reaksi hingga langkah aktivasi panas terjadi. Berbagai metode tersedia untuk menghentikan aktivitas hot start polymerase dan meliputi, modifikasi kimiawi, metode yang dimediasi antibodi, dan metode yang dimediasi aptamer. Endpoint PCR untuk Amplifikasi Target yang Panjang dan Akurat Endpoint PCR sering kali digunakan untuk mendeteksi keberadaan target dan kelimpahan relatif pada saat selesainya reaksi. Keterbatasan panjang sekuens yang dihasilkan selama PCR standar, sekitar 5 kb, sebagian diatasi dengan penggabungan faktor tambahan yang memberikan aktivitas "pengoreksian". PCR yang panjang dan akurat (LA) menggabungkan penggunaan polimerase termostabil kedua dengan eksonuklease 3′ → 5′ untuk memperbaiki kesalahan penggabungan nukleotida terminal, sehingga menghasilkan ketepatan yang meningkat secara signifikan dan kemampuan untuk mengamplifikasi target DNA hingga panjang 40 kb. [![Pencarian Dokumen](https://www.sigmaaldrich.com/content/dam/cms-commons/sigmaaldrich/marketing/global/images/ecommerce/document-search.png)](https://www.sigmaaldrich.com/documents-search) [Mencari Informasi yang Lebih Spesifik?](https://www.sigmaaldrich.com/documents-search) Kunjungi pencarian dokumen kami untuk mendapatkan lembar data, sertifikat, dan dokumentasi teknis. [Temukan Dokumen](https://www.sigmaaldrich.com/documents-search) ## Artikel Terkait - [Troubleshooting PCR and RT-PCR Amplification](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/pcr/troubleshooting-pcr-and-rt-pcr-amplification) Issues with PCR and RT-PCR are identified during gel electrophoresis analysis, such as the absence of bands, the presence of nonspecific bans, excessive smearing, and band smearing. - [Digital PCR](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/digital-pcr) Digital PCR enables absolute quantification and detection of minority sequences against similar majority sequences, such as somatic mutations. - [Ethidium Bromide](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/nucleic-acid-labeling-and-detection/ethidium-bromide) Ethidium bromide is a well-known and widely used fluorescent dye in biotechnology research. - [Polymerase Chain Reaction](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/pcr/polymerase-chain-reaction) Polymerase chain reaction is one of the most widely used techniques in molecular biology with 3 main steps: denature, anneal, and extend. - [Hot Start PCR](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/pcr/hot-start-pcr) The purpose of Hot Start PCR is to inhibit the PCR reaction in order to reduce nonspecific amplification, prevent the formation of primer dimers, and increase product yields. - [Lihat Semua (59)](https://www.sigmaaldrich.com/ID/id/search/facet-search?focus=sitecontent&term=facet-search) ## Protokol Terkait - [Standard PCR Protocol](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/standard-pcr) Learn standard PCR protocol steps and review reagent lists or cycling parameters. This method for routine PCR amplification of DNA uses standard Taq DNA polymerase. - [Annealing Oligonucleotides Protocol](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/annealing-oligos) Annealing is the process of heating and cooling two single-stranded oligonucleotides with complementary sequences. - [Oligonucleotide Melting Temperature](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/oligos-melting-temp) Learn about methods for calculating oligonucleotide melting temperature (Tm). - [PCR/qPCR/dPCR Assay Design](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/pcr-qpcr-dpcr-assay-design) PCR workflow's variability influenced by sample source, reverse transcription, and assay design, impacting reproducibility. - [REDExtract-N-Amp™ Tissue PCR Kit Protocol](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/redextract-n-amp) Extract-N-Amp kit protocol provides rapid DNA extraction yielding PCR-ready samples in 15 minutes with optimized PCR ReadyMix. - [Lihat Semua (59)](https://www.sigmaaldrich.com/ID/id/search/facet-search?focus=sitecontent&term=facet-search) ### Temukan Artikel dan Protokol Lainnya Masukkan Kata Kunci Pencarian ## Bagaimana Kami Dapat Membantu Jika ada pertanyaan, silakan kirimkan [permintaan dukungan pelanggan](https://www.sigmaaldrich.com/ID/id/support/customer-support) atau hubungi tim layanan pelanggan kami: Email [custserv@sial.com](mailto:custserv@sial.com) atau hubungi +1 (800) 244-1173 ## Dukungan Tambahan - [Chromatogram Search](https://www.sigmaaldrich.com/chromatogram-search) Use the Chromatogram Search to identify unknown compounds in your sample. - [Kalkulator & Aplikasi](https://www.sigmaaldrich.com/ID/id/support/calculators-and-apps) Web Toolbox - alat dan sumber daya penelitian ilmiah untuk kimia analitik, ilmu hayati, sintesis kimia, dan ilmu material. - [Customer Support Request](https://www.sigmaaldrich.com/ID/id/support/customer-support) Dukungan pelanggan termasuk bantuan dengan pesanan, produk, akun, dan masalah teknis situs web. - [FAQ](https://maestro.my.site.com/knowledgeportal/s/) Explore our Frequently Asked Questions for answers to commonly asked questions about our products and services. * * * __Artikel Pilihan__ [Troubleshooting PCR and RT-PCR Amplification](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/pcr/troubleshooting-pcr-and-rt-pcr-amplification) Issues with PCR and RT-PCR are identified during gel electrophoresis analysis, such as the absence of bands, the presence of nonspecific bans, excessive smearing, and band smearing. [Digital PCR](https://www.sigmaaldrich.com/ID/en/technical-documents/protocol/genomics/pcr/digital-pcr) Digital PCR enables absolute quantification and detection of minority sequences against similar majority sequences, such as somatic mutations. [Ethidium Bromide](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/nucleic-acid-labeling-and-detection/ethidium-bromide) Ethidium bromide is a well-known and widely used fluorescent dye in biotechnology research. [Polymerase Chain Reaction](https://www.sigmaaldrich.com/ID/en/technical-documents/technical-article/genomics/pcr/polymerase-chain-reaction) Polymerase chain reaction is one of the most widely used techniques in molecular biology with 3 main steps: denature, anneal, and extend. Atas __Masuk untuk Melanjutkan__ Untuk melanjutkan membaca, silakan masuk atau buat akun. Masuk__Tidak Punya Akun?__Daftar Untuk kenyamanan pelanggan kami, halaman ini telah diterjemahkan melalui terjemahan mesin. 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