{"id":33369,"date":"2025-11-13T16:01:50","date_gmt":"2025-11-13T21:01:50","guid":{"rendered":"https:\/\/genomequebec.wp.vortexdev.com\/en\/technological-services\/services-de-design-damorces-amplification-pcr-sequencage-sanger-et-analyse\/"},"modified":"2025-11-17T07:33:20","modified_gmt":"2025-11-17T12:33:20","slug":"primer-design-pcr-amplification-sanger-sequencing-and-analysis-services","status":"publish","type":"service","link":"https:\/\/genomequebec.wp.vortexdev.com\/en\/technological-services\/primer-design-pcr-amplification-sanger-sequencing-and-analysis-services\/","title":{"rendered":"Primer design, PCR amplification, Sanger sequencing and analysis services"},"content":{"rendered":"\n<h2 class=\"wp-block-heading has-text-align-center\" id=\"top-of-page\">3730xl DNA Analyzer Applied Biosystems technology<\/h2>\n\n\n\n<h3 class=\"wp-block-heading has-text-align-center has-secondary-color has-text-color has-link-color wp-elements-88d592e510ca54eee557f862b90f1bb2\"><em><strong><mark style=\"background-color:rgba(0, 0, 0, 0)\" class=\"has-inline-color has-primary-color\"><strong><em>User Guide<\/em><\/strong><\/mark><\/strong><\/em><\/h3>\n\n\n\n<p>The instructions in this guide must be followed carefully to achieve the best possible results and avoid delays in processing your request.<\/p>\n\n\n\n<p>Samples that do not comply with the recommendations may be rejected without compensation.<\/p>\n\n\n\n<p>Note that delays in the processing of samples will vary depending on the size of the project. It is recommended to contact the <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/contact-us\/\" target=\"_blank\" rel=\"noreferrer noopener\">Client Management Office<\/a> for more information.<\/p>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<ul class=\"wp-block-list\">\n<li><a href=\"#Starting-Material\" data-type=\"internal\" data-id=\"#Starting-Material\">Starting Material<\/a><\/li>\n\n\n\n<li><a href=\"#Requirements\" data-type=\"internal\" data-id=\"#Requirements\">Requirements<\/a><\/li>\n\n\n\n<li><a href=\"#Primers\" data-type=\"internal\" data-id=\"#Primers\">Primers<\/a>\n<ul class=\"wp-block-list\">\n<li class=\"has-black-color has-text-color has-link-color wp-elements-126f1a55da4f3e7a6cb305a78f713156\"><a href=\"#PCR-primer-to-design\" data-type=\"internal\" data-id=\"#PCR-primer-to-design\">PCR primer to design<\/a><\/li>\n\n\n\n<li class=\"has-black-color has-text-color has-link-color wp-elements-c5d246811f26d84664a62daa25483bbb\"><a href=\"#PCR-primers-whose-sequence-is-provided\" data-type=\"internal\" data-id=\"#PCR-primers-whose-sequence-is-provided\">PCR primers whose sequence is provided<\/a><\/li>\n\n\n\n<li class=\"has-black-color has-text-color has-link-color wp-elements-7f01c744934fa14725329d2643ad0628\"><a href=\"#PCR-and-or-sequencing-primers-provided\" data-type=\"internal\" data-id=\"#PCR-and-or-sequencing-primers-provided\">PCR and\/or sequencing primers provided<\/a><\/li>\n<\/ul>\n<\/li>\n\n\n\n<li><a href=\"#Required-containers-and-sample-identification\">Required containers and sample identification<\/a><\/li>\n\n\n\n<li><a href=\"#Service-Request-Form-and-Sample-Submission\" data-type=\"internal\" data-id=\"#Service-Request-Form-and-Sample-Submission\">Service Request Form and Sample Submission<\/a><\/li>\n\n\n\n<li><a href=\"#Sample-Shipment-Preparation\" data-type=\"internal\" data-id=\"#Sample-Shipment-Preparation\">Sample Shipment Preparation<\/a><\/li>\n\n\n\n<li><a href=\"#Transmission-of-results\" data-type=\"internal\" data-id=\"#Transmission-of-results\">Transmission of results<\/a><\/li>\n<\/ul>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-f1ea74b888da7196577b0d8ad8e41b53\" id=\"Starting-Material\">Starting Material<\/h4>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><thead><tr><th><strong><strong>Sample Type<\/strong><\/strong><\/th><th><strong>Preparation<\/strong><\/th><\/tr><\/thead><tbody><tr><td>Biological sample requiring extraction<\/td><td>Contact the <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/contact-us\/\" target=\"_blank\" rel=\"noreferrer noopener\">Client Management Office<\/a><br>for information on the <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/technological-services\/centre-dexpertise-et-de-services-2\/extraction\/\">Extraction Service<\/a><\/td><\/tr><tr><td>Genomic DNA (gDNA)<\/td><td><strong>Commercial kits<\/strong> recommended.<br><br><strong>In-house protocol<\/strong>: An additional purification step with a commercial kit is required to remove all traces of phenol\/chloroform.<br><br><strong>Elution<\/strong>: 10 mM Tris buffer, pH 8.5, or ultrapure water without nuclease.<\/td><\/tr><tr><td>Bacterial culture<\/td><td>Liquid bacterial culture (~8 hours)<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<div class=\"wp-block-buttons is-horizontal is-content-justification-center is-layout-flex wp-container-core-buttons-is-layout-03627597 wp-block-buttons-is-layout-flex\">\n<div class=\"wp-block-button\"><a class=\"wp-block-button__link wp-element-button\" href=\"#top-of-page\">Return to top of page<\/a><\/div>\n<\/div>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-a97d80a986aee5f5f1f68d86faa82c85\" id=\"Requirements\">Requirements<\/h4>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><thead><tr><th><strong><strong>Sample Type<\/strong><\/strong><\/th><th><strong>Volume<\/strong><\/th><th><strong>Concentration<\/strong><\/th><th><strong><strong>Recommended method for verifying concentration<\/strong><\/strong><\/th><th><strong><strong>Recommended method for verifying purity<\/strong><\/strong><\/th><th><strong><strong>Quality criteria to be met<\/strong><\/strong><\/th><\/tr><\/thead><tbody><tr><td>gDNA<\/td><td>10 \u00b5L of gDNA<br>per fragment to be analyzed<\/td><td>20 ng\/\u00b5L<\/td><td>Qubit or equivalent fluorometric method.<br><br><strong>Note<\/strong>: Spectrophotometric methods such as Nanodrop are much less accurate for measuring DNA concentration.<\/td><td>Nanodrop or other spectrophotometric method.<br><br><strong>Target optical density ratios:&nbsp;<\/strong><br><br>&#8211; 260\/280 between 1.8 and 2.0&nbsp;<br><br>&#8211; 260\/230 between 2.0 and 2.2.&nbsp;<\/td><td>No degraded or fragmented.<\/td><\/tr><tr><td>Bacterial culture<\/td><td>1 mL<\/td><td><strong>Target optical density ratios:<\/strong><br><br>&#8211; OD600 minimum: ~0.4 to 0.6 (growing bacteria)<br><br>&#8211; OD600 maximum: ~1.0 to 1.2 (to avoid overpopulation, which can induce cellular stress and reduce the quality of biomolecule)<\/td><td>Spectrophotometric<\/td><td>&nbsp;<\/td><td>&nbsp;<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<div class=\"wp-block-buttons is-horizontal is-content-justification-center is-layout-flex wp-container-core-buttons-is-layout-03627597 wp-block-buttons-is-layout-flex\">\n<div class=\"wp-block-button\"><a class=\"wp-block-button__link wp-element-button\" href=\"#top-of-page\">Return to top of page<\/a><\/div>\n<\/div>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-c617843e7d54b7a94d1b25c6791fd7a7\" id=\"Primers\">Primers<\/h4>\n\n\n\n<hr class=\"wp-block-separator has-alpha-channel-opacity is-style-default\" style=\"margin-top:24px;margin-bottom:24px\"\/>\n\n\n\n<h6 class=\"wp-block-heading has-blue-800-color has-text-color has-link-color wp-elements-0de8e2ccc91661550e296cf75d02775d\" id=\"PCR-primer-to-design\">PCR primer to design<\/h6>\n\n\n\n<p>When PCR primers are to be designed, provide the required information such as the rs (Reference SNP) number, the reference sequence, the position in the genome according to UCSC Genome Browser, the species, etc.<\/p>\n\n\n\n<div style=\"height:30px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<hr class=\"wp-block-separator has-alpha-channel-opacity\" style=\"margin-top:var(--wp--preset--spacing--60);margin-bottom:var(--wp--preset--spacing--60)\"\/>\n\n\n\n<h6 class=\"wp-block-heading has-blue-800-color has-text-color has-link-color wp-elements-6fb77b392a5eedbc5e55ed400566c215\" id=\"PCR-primers-whose-sequence-is-provided\">PCR primers whose sequence is provided<\/h6>\n\n\n\n<p>When the PCR primer sequence is provided by the customer Required containers and sample identification, indicate:<\/p>\n\n\n\n<p>\u2022 If these primers are to be ordered and if so, provide the name and sequence of each forward and reverse primer in the <a href=\"https:\/\/genomeqc.sharepoint.com\/:x:\/r\/sites\/Documentscontroles\/Documents%20partags_ChefsNanuqBGC\/Guides%20de%20l%27utilisateur%20pour%20site%20web%20GQ\/Amplicons%20-%20Sanger_Formulaire%20de%20soumisison%20d%27%C3%A9chantillons-Sample%20submission%20form_FR_EN.xlsx?d=w54d056a29e124f56a6dc79811b599a4a&amp;csf=1&amp;web=1&amp;e=PykqKT\" target=\"_blank\" rel=\"noreferrer noopener\">Sample Submission Form<\/a> in the corresponding tab.<\/p>\n\n\n\n<p>\u2022 If they are part of the standard primers offered free of charge by the PCR amplification \u2013 Sanger Sequencing service, see the following list:<\/p>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><tbody><tr><td>27F<\/td><td>5\u2019 \u2013 AGAGTTTGATCMTGGCTCAG &#8211; 3\u2019<\/td><\/tr><tr><td>1492R<\/td><td>5\u2019 \u2013 TACGGYTACCTTGTTACGACTT &#8211; 3\u2019<\/td><\/tr><tr><td>ITS1F<\/td><td>5&#8242;-CTTGGTCATTTAGAGGAAGTAA-3&#8242;<\/td><\/tr><tr><td>ITS4<\/td><td>5&#8242;-TCCTCCGCTTATTGATATGC-3&#8242;<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<hr class=\"wp-block-separator has-alpha-channel-opacity\" style=\"margin-top:var(--wp--preset--spacing--60);margin-bottom:var(--wp--preset--spacing--60)\"\/>\n\n\n\n<h6 class=\"wp-block-heading has-blue-800-color has-text-color has-link-color wp-elements-1cf55ed4133f6ea5a807198d38ee0fd7\" id=\"PCR-and-or-sequencing-primers-provided\">PCR and\/or sequencing primers provided<\/h6>\n\n\n\n<p>When PCR and\/or sequencing primers are provided by the customer, the following rules must be observed:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>The primers must not contain any known SNPs. For any clarification, contact the <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/contact-us\/\" target=\"_blank\" rel=\"noreferrer noopener\">Client Management Office<\/a>.<\/li>\n\n\n\n<li>The primer length must be be between 18 and 24 bases.<\/li>\n\n\n\n<li>The G\/C ratio should be 40-60%.<\/li>\n\n\n\n<li>PCR product size should be between 250 and 750 bases.<\/li>\n\n\n\n<li>The primer annealing temperature (Tm) must be above 50\u00b0C.<\/li>\n\n\n\n<li>Primers must be at least 100 bases from the target region to be sequenced.<\/li>\n\n\n\n<li>Primer design upstream of homo- or heteropolymeric regions (repeats of A, C, G, or T) should be avoided, as these regions are extremely difficult to sequence.<\/li>\n\n\n\n<li>The Tm and %GC of the primers must be provided.<\/li>\n<\/ul>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><thead><tr><th class=\"has-text-align-center\" data-align=\"center\"><strong><strong>Sample Type<\/strong><\/strong><\/th><th class=\"has-text-align-center\" data-align=\"center\"><strong>Volume<\/strong><\/th><th class=\"has-text-align-center\" data-align=\"center\"><strong>Concentration<\/strong><\/th><\/tr><\/thead><tbody><tr><td class=\"has-text-align-center\" data-align=\"center\">Primer<\/td><td class=\"has-text-align-center\" data-align=\"center\">5 \u00b5L <br>per sample to be analyzed<\/td><td class=\"has-text-align-center\" data-align=\"center\">20 \u00b5M<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<div class=\"wp-block-buttons is-horizontal is-content-justification-center is-layout-flex wp-container-core-buttons-is-layout-03627597 wp-block-buttons-is-layout-flex\">\n<div class=\"wp-block-button\"><a class=\"wp-block-button__link wp-element-button\" href=\"#top-of-page\">Return to top of page<\/a><\/div>\n<\/div>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-9375746457abecc41d782c59deaf1249\" id=\"Required-containers-and-sample-identification\"><strong>Required containers and sample identification<\/strong><\/h4>\n\n\n\n<ul class=\"wp-block-list\">\n<li><strong>48 samples or less<\/strong>: &#8220;strip&#8221; tubes or 96-well PCR plate. <\/li>\n\n\n\n<li><strong>48 samples or more<\/strong>: 96-well PCR plate.<\/li>\n<\/ul>\n\n\n\n<p>The name on the tube or plate must be unique,&nbsp;readable, concise, and identical to the name&nbsp;indicated&nbsp;on the submission form.&nbsp;<\/p>\n\n\n\n<p>Samples must be placed on the plate as indicated on the submission form &#8211; <span style=\"text-decoration: underline;\">Row arrangement mandatory<\/span>.<\/p>\n\n\n\n<p>This is to avoid any delays in processing the request.<\/p>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><thead><tr><th><strong><strong><strong>Sample Type<\/strong><\/strong><\/strong><\/th><th><strong><strong>Container required<\/strong><\/strong><\/th><th><strong><strong>Recommended containers<\/strong><\/strong><\/th><th><strong><strong>Containers refused<\/strong><\/strong><\/th><\/tr><\/thead><tbody><tr><td>Bacterial culture<\/td><td>&#8211; 1.5mL tubes<br>&nbsp;<br>All types of 1.5 mL tubes with attached snap caps are accepted.<br><br>Example: <em>DNA LoBind Tubes<\/em>, 1.5 mL, <em>PCR clean, colorless<\/em>; Eppendorf, Cat # 022431021<\/td><td>&nbsp;<\/td><td>&nbsp;<\/td><\/tr><tr><td>gDNA<\/td><td>&nbsp;<\/td><td>&#8211; PCR strip tubes of 8 or 12<br><br>All types of 200\u00b5L PCR tubes in strip of 8 or 12 with a cap are accepted.<br><br>Example: <em>0,2 mL PCR strip tubes with 12 well<\/em>, VWR, Cat # 53509-300\u00a0<br><br>With <em>strip domed caps for 12 well strips<\/em>, VWR, Cat # 53509-302\u00a0\u00a0 \u00a0 <br><br>&#8211; Half-skirt PCR plate<br><br>All types of\u00a0clear\u00a0half-skirt clear 96-well PCR plates\u00a0are accepted.\u00a0<br><br>Example: <em>Thermo-Fast 96 PCR detection plate with flat deck<\/em>; Life Technologie, Cat# AB1400L<br><br>Exemple: <em>Thermo-Fast 96 PCR detection plate with flat deck<\/em>; Life Technologie, Cat# AB1400L <br><br>&#8211; Full-skirt\u00a0PCR plate\u00a0\u00a0<br><br>All types of clear\u00a0full-skirt 96-well PCR plates\u00a0are accepted.\u00a0<br><br>&#8211; Example: <em>Microseal\u00a0PCR plates 96-well clear; Bio-Rad, Cat#\u202fMSP9601<\/em>\u00a0<br><br><strong><strong>Clear adhesive film<\/strong><br><strong>recommended:<\/strong><\/strong><br><br>&#8211; Clear adhesive film (Thermo Fisher Scientific, Cat# AB-0558)<br><br>&#8211; 8 or 12-strip caps ensuring a tight seal<br><br>&#8211; Heat sealing to prevent leaks or cross-contamination<\/td><td>&#8211; 96-well cell culture plates<br><br>&#8211; No-skirt 96-well PCR plates<br><br>&#8211; Opaques 96-well PCR plates<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<div class=\"wp-block-buttons is-horizontal is-content-justification-center is-layout-flex wp-container-core-buttons-is-layout-03627597 wp-block-buttons-is-layout-flex\">\n<div class=\"wp-block-button\"><a class=\"wp-block-button__link wp-element-button\" href=\"#top-of-page\">Return to top of page<\/a><\/div>\n<\/div>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-1afbfc3d09342a442ee7af6d6f5edfd7\" id=\"Service-Request-Form-and-Sample-Submission\">Service Request Form and Sample Submission<\/h4>\n\n\n\n<p>Click on <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/technological-services\/centre-dexpertise-et-de-services-2\/instructions\/service-request-and-sample-submission\/\" target=\"_blank\" rel=\"noreferrer noopener\">service request and sample submission<\/a> to view instructions.&nbsp;<\/p>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-096fea86e9448c1a40045488fbf1d398\" id=\"Sample-Shipment-Preparation\">Sample Shipment Preparation<\/h4>\n\n\n\n<p>Click on <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/technological-services\/centre-dexpertise-et-de-services-2\/instructions\/sample-shipment-preparation\/\" target=\"_blank\" rel=\"noreferrer noopener\">samples shipment preparation<\/a> to view instructions.&nbsp;<\/p>\n\n\n\n<div style=\"height:35px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<h4 class=\"wp-block-heading has-secondary-color has-text-color has-link-color wp-elements-001b4d0493cf1fc677438e86372a3251\" id=\"Transmission-of-results\">Transmission of results<\/h4>\n\n\n\n<p>Sequencing and analysis results, if applicable, are directly accessible online in <a href=\"https:\/\/ces.genomequebec.com\/nanuqAdministration\/j_security_check?service=http%3A%2F%2Fces.genomequebec.com%2FnanuqAdministration%2F\" target=\"_blank\" rel=\"noreferrer noopener\">Nanuq<\/a>.<\/p>\n\n\n\n<p>An email is sent at the end of the project to inform of the availability of the results.<\/p>\n\n\n\n<p>\u2714\ufe0f Chromatograms and text sequences (FASTA or GenBank format) can be viewed and downloaded locally.<\/p>\n\n\n\n<p>\u2714\ufe0f The SNP detection analysis report contains the following information:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>The position of the SNPs found in the genome<\/li>\n\n\n\n<li>The identification number (rs) if the SNP is already listed<\/li>\n\n\n\n<li>The genotype of each sample analyzed<\/li>\n\n\n\n<li>Whether the SNP is intronic or exonic, and if so, whether it is a synonymous or non-synonymous SNP<\/li>\n\n\n\n<li>The amino acid change, if applicable.<\/li>\n\n\n\n<li>The Excel report contains a tab with a tutorial to assist in data interpretation.<\/li>\n<\/ul>\n\n\n\n<p>\u2714\ufe0f The Excel genotyping analysis report contains the genotype of each sample analyzed.<\/p>\n\n\n\n<p>\u2714\ufe0f The BLAST analysis Word report contains the species and genus, E-value, percent identity, and accession number.<\/p>\n\n\n\n<p>For any questions, please contact the <a href=\"https:\/\/genomequebec.wp.vortexdev.com\/en\/contact-us\/\" target=\"_blank\" rel=\"noreferrer noopener\">Client Management Office<\/a> at any time.<\/p>\n\n\n\n<p><strong>Version 1.0_2025-11-06<\/strong><\/p>\n\n\n\n<div class=\"wp-block-buttons is-horizontal is-content-justification-center is-layout-flex wp-container-core-buttons-is-layout-03627597 wp-block-buttons-is-layout-flex\">\n<div class=\"wp-block-button\"><a class=\"wp-block-button__link wp-element-button\" href=\"#top-of-page\">Return to top of page<\/a><\/div>\n<\/div>\n","protected":false},"featured_media":0,"parent":0,"menu_order":0,"template":"","class_list":["post-33369","service","type-service","status-publish","hentry"],"acf":[],"_links":{"self":[{"href":"https:\/\/genomequebec.wp.vortexdev.com\/en\/wp-json\/wp\/v2\/service\/33369","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/genomequebec.wp.vortexdev.com\/en\/wp-json\/wp\/v2\/service"}],"about":[{"href":"https:\/\/genomequebec.wp.vortexdev.com\/en\/wp-json\/wp\/v2\/types\/service"}],"wp:attachment":[{"href":"https:\/\/genomequebec.wp.vortexdev.com\/en\/wp-json\/wp\/v2\/media?parent=33369"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}