{ "cells": [ { "cell_type": "markdown", "metadata": { "colab_type": "text", "id": "XBwlMr3_KXD2" }, "source": [ "# Case Study: Thermal Cycling for PCR\n", "\n", "The purpose of this notebook is to describe a technological challenge requiring the use of the feedback control for a satisfactory result. Here we consider performance requirements for a thermal cycling device used for diagnostic testing based on the polymerase chain reaction (PCR). These devices are used to exponentially \"amplify\" segments of DNA that may be present in biological samples. These devices are ubiquitous in modern biology laboraties and medical testing services.\n", "\n", "In this notebook we will review:\n", "\n", "* Polymerase chain reaction (PCR) \n", "* Several PCR thermal cyclers used in diagnostic laboratories\n", "* Performance requirements for temperature control in a PCR thermal cycler\n", "* Use of Temperature Control Laboratory to prototype a PCR control system\n", "\n", "The learning goals are for student to able to:\n", "\n", "* Describe use the of feedback control as an enabling technology for laboratory and process systems.\n", "* Develop control performance requirements for an application." ] }, { "cell_type": "markdown", "metadata": { "colab_type": "text", "id": "2IaCjA9tN9gy" }, "source": [ "## Pre-reads\n", "\n", "* [Notre Dame Center for Advanced Diagnostics and Therapeutics](https://advanceddiagnostics.nd.edu/assets/382241/coronavirus_test_faq.pdf)\n", "* [miniPCR Thermal Cycler]( https://www.youtube.com/watch?time_continue=1&v=ALNZJhUOSMs&feature=emb_logo)" ] }, { "cell_type": "markdown", "metadata": { "colab_type": "text", "id": "ta4O_dtbaBah" }, "source": [ "## Coronavirus Diagnostics\n", "\n", "COVID-19 is a respiratory tract infection by a specific species of coronavirus called SARS-CoV-2. Coronaviruses have a protein envelope characterized by club-shaped protrusions that give the impression of a corona when viewed with an electron microscope.\n", "\n", "

\"Electron
By NIAID - https://www.flickr.com/photos/niaid/49534865371/, CC BY 2.0, Link

\n", "\n", "Coronaviruses have a relatively large genome comprised of a single strand of positive-sense RNA.\n", "\n", "

\"Vaccines-08-00587-g002-A\"

\n", "\n", "[https://commons.wikimedia.org/wiki/File:Vaccines-08-00587-g002-A.png](https://commons.wikimedia.org/wiki/File:Vaccines-08-00587-g002-A.png)\n", "\n", " A coronavirus test begins with a swab of throat or nose. The swab is rinsed with a buffer solution to capture the host DNA and RNA, and the RNA of any virus that may be present. RNA is isolated from the solution and converted to DNA using a reverse transcriptase enzyme. The amount of DNA collected in this fashion is too small to analyze by conventional methods. So the first step in a diagnostic protocol is to amplify the amount of DNA using the polymerase chain reaction." ] }, { "cell_type": "markdown", "metadata": { "colab_type": "text", "id": "AEQ15Fd8N7iR" }, "source": [ "## Polymerase Chain Reaction (PCR)\n", "\n", "The polymerase chain reaction (PCR) is a technique used in molecular biology to take amplify small samples of DNA into quantities large enough to be detected using conventional analytical methods. The technique is described in the following videos." ] }, { "cell_type": "code", "execution_count": 1, "metadata": {}, "outputs": [ { "data": { "image/jpeg": 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"text/html": [ "\n", " \n", " " ], "text/plain": [ "" ] }, "execution_count": 1, "metadata": {}, "output_type": "execute_result" } ], "source": [ "from IPython.lib.display import YouTubeVideo\n", "YouTubeVideo('2KoLnIwoZKU')" ] }, { "cell_type": "code", "execution_count": 2, "metadata": {}, "outputs": [ { "data": { "image/jpeg": 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", "text/html": [ "\n", " \n", " " ], "text/plain": [ "" ] }, "execution_count": 2, "metadata": {}, "output_type": "execute_result" } ], "source": [ "from IPython.lib.display import YouTubeVideo\n", "YouTubeVideo('c07_5BfIDTw')" ] }, { "cell_type": "markdown", "metadata": {}, "source": [ "PCR provides an exponential amplification of target DNA sequences to concentrations that can be detected by conventional analytical techniques.\n", "\n", "\"Polymerase\n", "\n", "[https://commons.wikimedia.org/wiki/File:Polymerase_chain_reaction-en.svg](https://commons.wikimedia.org/wiki/File:Polymerase_chain_reaction-en.svg)\n", "\n", "The main part of PCR consists of three steps that are repeated 20 or more times:\n", "\n", "1. Denaturation at 94-96 $^\\circ$C. At this step, DNA 'breaks apart', splitting from a double helix into single strands\n", "2. Annealing at temperatures between 50 and 68 $^\\circ$C. Primers bond to the single-stranded DNA. The exact temperature depends on the length and content of the target sequence.\n", "3. Extension at ca. 72 $^\\circ$C. Polymerase compliments the DNA, synthesizing strands that are of the target sequence" ] }, { "cell_type": "markdown", "metadata": { "colab_type": "text", "id": "EjJ1NaKvNYzz" }, "source": [ "## Examples of low-cost and open-source PCR thermal cyclers\n", "\n", "PCR thermal cyclers are a common piece of hardware in biology and medical testing laboratories. These devices automate the thermal cycle operation by providing users with convenient means of specifying parameters for a particular PCR test, and then executing that cycle to some specified degree of precision. Most commercial devices perform multiple from a few to several hundres tests at a time.\n", "\n", "![](http://media.biocompare.com/m/37/Product/239450-400x300.jpg)\n", "\n", "There are also examples of low-cost and open-source designs of PCR thermal cyclers intended for education, field work, and other situations where there is a benefit to keeping costs very low.\n", "\n", "* [miniPCR Thermal Cycler]( https://www.youtube.com/watch?time_continue=1&v=ALNZJhUOSMs&feature=emb_logo)\n", "* https://www.instructables.com/id/Arduino-PCR-thermal-cycler-for-under-85/\n" ] }, { "cell_type": "code", "execution_count": 3, "metadata": { "colab": { "base_uri": "https://localhost:8080/", "height": 321 }, "colab_type": "code", "executionInfo": { "elapsed": 1405, "status": "ok", "timestamp": 1584913120241, "user": { "displayName": "Jeffrey Kantor", "photoUrl": "https://lh3.googleusercontent.com/a-/AOh14Gg_n8V7bVINy02QRuRgOoMo11Ri7NKU3OUKdC1bkQ=s64", "userId": "09038942003589296665" }, "user_tz": 240 }, "id": "scK7j3pTKNVT", "outputId": "e1b05aa1-835e-4e12-9f62-57f3196f0a94" }, "outputs": [ { "data": { "image/jpeg": 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", "text/html": [ "\n", " \n", " " ], "text/plain": [ "" ] }, "execution_count": 3, "metadata": {}, "output_type": "execute_result" } ], "source": [ "from IPython.display import YouTubeVideo\n", "YouTubeVideo('ALNZJhUOSMs')" ] }, { "cell_type": "markdown", "metadata": {}, "source": [ "## The Challenge: Prototyping a Control System for a PCR Thermal Cycler\n", "\n", "With two heater/sensor assemblies, the temperature Control Lab provides a platform for developing and testing control strategies for a PCR thermal cycler.\n", "\n", "\n", "\n", "There are some differences. PCR thermal cyclers generally include thermoelectric coolers rather than rely solely on convective cooling, and generally have more \"control authority\" in the form of larger power inputs. So we will be limited in our ability to mimic higher performance devices. But for the purposes of prototyping, the Temperature Contol Lab offers an excellent platform for initial prototype." ] }, { "cell_type": "markdown", "metadata": {}, "source": [ "## What will this project entail?\n", "\n", "![](../figures/B.01-FeedbackControlDiagram.png)\n", "\n", "Consider the use of the Temperature Control Lab as a prototyping device for a PCR Thermal Cycling device. Discuss each of the following aspects of the control loop:\n", "\n", "1. What are the control variables (CV)?\n", "2. What are the measurement process variables (PV)? Are they different than the CV's?\n", "3. What are the setpoints (SP)?\n", "4. What are the manipulated variables (MV)?\n", "5. What are some examples of disturbance variables (DV)? (There are many possibilities.)\n", "6. What diagnostics would be useful in this application?\n", "\n", "In qualitative terms, describe:\n", "\n", "1. Why is feedback control important for this application? \n", "2. Is there a role for feedforward control?\n", "3. How will we deal with the fact the CV's are not the same as the PV's?\n", "4. Describe some possible control algorithms that would allow you to track a setpoint.\n" ] }, { "cell_type": "code", "execution_count": null, "metadata": {}, "outputs": [], "source": [] } ], "metadata": { "colab": { "authorship_tag": "ABX9TyNMv0PCsIVk9+5F36a4xQRI", "collapsed_sections": [], "name": "01.00-PCR_Overview.ipynb", "provenance": [] }, "kernelspec": { "display_name": "Python 3 (ipykernel)", "language": "python", "name": "python3" }, "language_info": { "codemirror_mode": { "name": "ipython", "version": 3 }, "file_extension": ".py", "mimetype": "text/x-python", "name": "python", "nbconvert_exporter": "python", "pygments_lexer": "ipython3", "version": "3.9.13" } }, "nbformat": 4, "nbformat_minor": 4 }