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scinote-web/app/assets/templates/experiment_433/experiment.json
Andrej 230aea15fc Update experiment templates [SCI-7314]
2022-10-11 15:55:56 +02:00

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{
"experiment": {
"id": 433,
"name": "Development of SNP markers panel",
"description": "This template shows how to generate a SNP markers panel with the preparation of SNP panel.\r\nSingle nucleotide polymorphisms (SNPs) markers are widely used for genomic research and breeding. Development of a panel of diagnostic SNP markers is used for the genotyping as well as the identification of hybrids and interspecies introgression events.",
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"updated_at": "2019-02-07T14:44:00.337Z",
"uuid": "de7f0d1b-f842-440b-a367-a0333a11430c"
},
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"name": "RAD library preparation and sequencing",
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body>\n<p>PCR conditions: </p>\n<ul>\n<li>95°C for 1 min</li>\n<li>35 cycles: 95°C for 15 s</li>\n<li>56°C for 15 s</li>\n<li>72°C for 22 s</li>\n</ul>\n<br>PCR products run at <strong>60 V</strong> for <strong>40 mins</strong> on a<strong> 2%</strong> agarose gel (<strong>0.5× TAE</strong>, stained with <strong>100 ng/μL</strong> EtBr).</body></html>"
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>Libraries can be quantified by fluorimetry and calibrated by sequencing. </p></body></html>"
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"text": "120 μL of each amplified library is size-selected (about 250 to 500 bp) by gel electrophoresis.",
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"text": "Final libraries are put through quality control and high-throughput sequencing.",
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body>\n<p> The RAD library was prepared as originally described in <a href=\"https://www.ncbi.nlm.nih.gov/pubmed/18852878/\" target=\"_blank\" rel=\"noopener\">article</a>.<br><br>At least <strong>30</strong> specimens need to be chosen.<br><br><strong>Each sample is digested at 37°C for 40 min with restriction enzyme:</strong></p>\n<ul>\n<li>\n<strong>6 U</strong> <em>restriction enzyme</em> per μg genomic DNA in <strong>1×</strong> Reaction Buffer 4 at a final concentration of about <strong>1 μg</strong> DNA per <strong>50 μL</strong> reaction volume</li>\n<li>The samples are heat-inactivated at <strong>65°C</strong> for <strong>20 min</strong>.</li>\n</ul>\n<br><strong>Individual-specific P1 adapters, each with a unique 5 or 7 bp barcode is ligated to the digested DNA at 22°C for 15 min by adding:</strong><br><br>\n<ul>\n<li>0.6 μL (DNA samples) 100 nmol/L P1 adapter</li>\n<li>0.15 μL 100 mmol/L rATP </li>\n<li>0.25 μL 10× Reaction Buffer 2</li>\n<li>0.125 μL T4 ligase </li>\n<li>reaction volumes made up to 15 μL with nuclease-free water </li>\n</ul>\n<br>After heat-inactivation, the ligation reactions are slowly cooled to room temperature then combined in appropriate multiplex pools.</body></html>"
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body>\n<p>Reads of low quality must be discarded. <br>Retained reads are sorted into loci and genotypes using software developed for that purpose. It will assigns loci using a likelihood-based algorithm to separate actual SNPs from SNPs likely to have arisen from sequencing error. <br><br><strong>Parameters:</strong><br>\n</p>\n<ul>\n<li>De novo assembly parameters.</li>\n<li>Minimum stack depth of 5.</li>\n<li>Maximum of 2 mismatches.</li>\n<li>No more than 1 mismatch between alleles.</li>\n</ul>\n</body></html>"
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>Sequencing data from filtered Informative RAD markers is combined into a single alignment of alleles in RAD library construction. Data analysis is carried out using R and an associated R/<em>adegenet</em> package for Principal Component Analysis (PCA) and Discriminant Analysis of Principal Components (DAPC).<em><strong> PCA creates simplified models of the total variation within the dataset.</strong> <strong>DAPC identifies clusters of genetically related individuals.</strong></em></p></body></html>"
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>Reading the fluorescence emission of the FAM and HEX fluorophores for each sample, in comparison to no-template control reactions, using a Real Time PCR Thermal Cycler and Quansoft endpoint genotyping software.</p></body></html>"
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"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body>\n<p>SNP of interest at a given locus need to be at least <strong>20 bp</strong> from the end of a given sequence, to allow for primer design. SNP assays were designed and manufactured for use with KASP genotyping technology.<br><br><strong>Optimisation assay conditions:</strong></p>\n<ul>\n<li>94°C for 15 min;</li>\n<li>10 cycles 94°C for 20 s, 61–55°C for 120 s (0.6°C drop per cycle)</li>\n<li>40 cycles 94°C for 20 s, 55°C for 120 s </li>\n</ul>\n</body></html>"
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"created_at": "2018-11-12T12:00:57.856Z",
"updated_at": "2018-12-24T08:46:24.129Z",
"last_modified_by_id": 202,
"protocol_id": 3487
},
"step_comments": [
],
"assets": [
],
"step_orderable_elements": [
{
"step_text": {
"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>Here are the guidelines for protein extraction: </p></body></html>"
},
"position": 0
},
{
"checklist": {
"checklist": {
"id": 958,
"name": "Guidelines:",
"step_id": 4416,
"created_at": "2018-12-21T09:50:59.139Z",
"updated_at": "2018-12-21T09:50:59.139Z",
"created_by_id": null,
"last_modified_by_id": null
},
"checklist_items": [
{
"id": 3939,
"text": "Add 1 volume of chloroform: Isoamyl alcohol to the solution and mix by inversion for 5 min. Centrifuge the sample for 10 min at 5000 × g and pipette the upper aqueous phase into a new Falcon tube.",
"checked": false,
"checklist_id": 958,
"created_at": "2018-12-21T09:50:59.141Z",
"updated_at": "2018-12-21T09:50:59.141Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 0
},
{
"id": 3940,
"text": "Add 5 μL of RNAse A to the solution and incubate at 37°C for 15 min with periodic, gentle mixing.",
"checked": false,
"checklist_id": 958,
"created_at": "2018-12-21T09:50:59.149Z",
"updated_at": "2018-12-21T09:50:59.149Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 1
},
{
"id": 3941,
"text": "After incubation, add 1 volume of chloroform: Isoamyl alcohol to the solution and mix by inversion for 5 min.",
"checked": false,
"checklist_id": 958,
"created_at": "2018-12-21T09:50:59.156Z",
"updated_at": "2018-12-21T09:50:59.156Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 2
},
{
"id": 3942,
"text": "Centrifuge the solution for 10 min at 5000 × g and pipette the aqueous phase into a new Falcon tube.",
"checked": false,
"checklist_id": 958,
"created_at": "2018-12-21T09:50:59.163Z",
"updated_at": "2018-12-21T09:50:59.163Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 3
}
]
},
"position": 1
}
]
},
{
"step": {
"id": 4418,
"name": "DNA quality and quantity assessment",
"position": 3,
"completed": false,
"completed_on": null,
"user_id": 202,
"created_at": "2018-11-12T12:22:47.725Z",
"updated_at": "2019-02-19T14:34:47.503Z",
"last_modified_by_id": 202,
"protocol_id": 3487
},
"step_comments": [
],
"assets": [
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"step_orderable_elements": [
{
"step_text": {
"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body>\n<p>Looking for a single absorbance peak at <strong>260 nm</strong>, a <strong>260/280</strong> absorbance ratio of <strong>1.8-2.0</strong> and no evidence of substantial band shearing or contamination (either RNA or polysaccharide).<br><br></p>\n<p><strong>Assess the quality of the extracted DNA:</strong></p>\n<ol>\n<li>NanoDrop UV/Vis spectrophotometer</li>\n<li>0.7% (w/v) agarose gel electrophoresis</li>\n</ol>\n</body></html>"
},
"position": 0
}
]
},
{
"step": {
"id": 4414,
"name": "Preparation",
"position": 0,
"completed": false,
"completed_on": null,
"user_id": 202,
"created_at": "2018-11-12T11:06:48.312Z",
"updated_at": "2018-12-24T08:34:20.404Z",
"last_modified_by_id": 202,
"protocol_id": 3487
},
"step_comments": [
],
"assets": [
],
"step_orderable_elements": [
{
"step_text": {
"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>Guidelines for samples preparation below. </p></body></html>"
},
"position": 0
},
{
"checklist": {
"checklist": {
"id": 957,
"name": "Guidelines",
"step_id": 4414,
"created_at": "2018-12-21T09:43:55.386Z",
"updated_at": "2018-12-21T09:43:55.386Z",
"created_by_id": null,
"last_modified_by_id": null
},
"checklist_items": [
{
"id": 3934,
"text": "Samples are stored in 95% ethanol solution at -20°C.",
"checked": false,
"checklist_id": 957,
"created_at": "2018-12-21T09:43:55.389Z",
"updated_at": "2018-12-21T09:43:55.389Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 0
},
{
"id": 3935,
"text": "Pre-heat water baths (65°C and 37°C).",
"checked": false,
"checklist_id": 957,
"created_at": "2018-12-21T09:43:55.396Z",
"updated_at": "2018-12-21T09:43:55.396Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 1
},
{
"id": 3936,
"text": "Prepare 10 mL (per 1 g of sample) extraction buffer by adding 0.3% (v/v) β-mercaptoethanol in a 50 mL Falcon tube, and pre-heat in the 65°C water bath.",
"checked": false,
"checklist_id": 957,
"created_at": "2018-12-21T09:43:55.403Z",
"updated_at": "2018-12-21T09:43:55.403Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 2
},
{
"id": 3937,
"text": "Put the sample into the 65°C water bath and mix by inversion every 10 min for 30-60 minutes.",
"checked": false,
"checklist_id": 957,
"created_at": "2018-12-21T09:43:55.412Z",
"updated_at": "2018-12-21T09:43:55.412Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 3
},
{
"id": 3938,
"text": "After incubation, centrifuge the sample tube for 5 min at 5000 × g and decant the supernatant into a new 50 mL Falcon tube.",
"checked": false,
"checklist_id": 957,
"created_at": "2018-12-21T09:43:55.419Z",
"updated_at": "2018-12-21T09:43:55.419Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 4
}
]
},
"position": 1
}
]
},
{
"step": {
"id": 4417,
"name": "Precipitation",
"position": 2,
"completed": false,
"completed_on": null,
"user_id": 202,
"created_at": "2018-11-12T12:05:42.004Z",
"updated_at": "2019-02-07T10:03:24.266Z",
"last_modified_by_id": 202,
"protocol_id": 3487
},
"step_comments": [
],
"assets": [
],
"step_orderable_elements": [
{
"step_text": {
"text": "<!DOCTYPE html PUBLIC \"-//W3C//DTD HTML 4.0 Transitional//EN\" \"http://www.w3.org/TR/REC-html40/loose.dtd\">\n<html><body><p>If you are precipitating small volumes of DNA, and you can fit the required amount of solvent into the sample tube, then ice-cold ethanol is the preferred choice. You can chill it (in liquid nitrogen or at <strong>–80°C</strong>) to accelerate the precipitation without the risk of precipitating excess salt. Afterwards, you need to wash the pellet with <strong>70%</strong> ethanol to remove any salt present.</p></body></html>"
},
"position": 0
},
{
"checklist": {
"checklist": {
"id": 959,
"name": "Guidelines:",
"step_id": 4417,
"created_at": "2018-12-21T09:54:21.494Z",
"updated_at": "2018-12-21T09:54:21.494Z",
"created_by_id": null,
"last_modified_by_id": null
},
"checklist_items": [
{
"id": 3943,
"text": "Add a ½ volume of 5 M NaCl to the sample and mix gently by inversion.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.496Z",
"updated_at": "2018-12-21T09:54:21.496Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 0
},
{
"id": 3944,
"text": "Then, add 3 volumes of cold 95% ethanol and mix gently by inversion.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.503Z",
"updated_at": "2018-12-21T09:54:21.503Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 1
},
{
"id": 3945,
"text": "Place the tubes into a -20°C freezer and incubate for 1 h.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.509Z",
"updated_at": "2018-12-21T09:54:21.509Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 2
},
{
"id": 3946,
"text": "After incubation, centrifuge the Falcon tube for 10 min at 5000 × g to pellet the DNA.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.516Z",
"updated_at": "2018-12-21T09:54:21.516Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 3
},
{
"id": 3947,
"text": "Carefully decant away the supernatant and wash the DNA pellet with 3 mL of 70% ethanol.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.522Z",
"updated_at": "2018-12-21T09:54:21.522Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 4
},
{
"id": 3948,
"text": "Gently swirl the solution and centrifuge again for 10 min at 5000 × g.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.529Z",
"updated_at": "2018-12-21T09:54:21.529Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 5
},
{
"id": 3949,
"text": "Carefully decant the supernatant and air-dry DNA pellet for 15 min at room temperature. Once dried, suspend DNA in 200 μL of TE buffer.",
"checked": false,
"checklist_id": 959,
"created_at": "2018-12-21T09:54:21.535Z",
"updated_at": "2018-12-21T09:54:21.535Z",
"created_by_id": null,
"last_modified_by_id": null,
"position": 6
}
]
},
"position": 1
}
]
}
]
}
],
"results": [
]
}
],
"my_module_groups": [
{
"id": 1167,
"created_at": "2018-12-17T07:51:25.142Z",
"updated_at": "2018-12-17T07:51:25.142Z",
"created_by_id": 202,
"experiment_id": 433
}
]
}
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