<html><head><meta http-equiv="Content-Type" content="text/html; charset=utf-8"></head><body style="word-wrap: break-word; -webkit-nbsp-mode: space; line-break: after-white-space;" class="">Hi Margaret,<div class=""><br class=""></div><div class="">The VEP cache files and the database both contain human SIFT and Polyphen data, so there should be no need to use this plugin. If you add the options '—sift b’ and/or ‘—polyphen b’ to your VEP command then you should receive the correct annotations (see <a href="https://www.ensembl.org/info/docs/tools/vep/script/vep_options.html#opt_sift" class="">https://www.ensembl.org/info/docs/tools/vep/script/vep_options.html#opt_sift</a>) </div><div class=""><br class=""></div><div class="">Regarding your connection issues to the ensembl database servers, are you able to successfully ping the ensembl servers with: </div><div class=""><pre style="font-family: "Courier New", Courier, monospace; font-size: 12.8px; margin-top: 0px; margin-bottom: 16px; line-height: 16px; overflow-wrap: break-word; color: rgb(102, 102, 102); font-variant-ligatures: normal; orphans: 2; widows: 2; background-color: rgb(255, 255, 255);" class="">ping -c 5 <a href="http://ensembldb.ensembl.org" class="">ensembldb.ensembl.org</a></pre><div class="">If this ping is successful, then it may be that port 3306 is not open on your network.</div></div><div class=""><br class=""></div><div class="">Kind Regards,</div><div class="">Andrew</div><div class=""><br class=""></div><div class=""><br class=""><div><blockquote type="cite" class=""><div class="">On 10 Mar 2020, at 23:24, Linan, Margaret <<a href="mailto:margaret.linan@mssm.edu" class="">margaret.linan@mssm.edu</a>> wrote:</div><br class="Apple-interchange-newline"><div class=""><div class="WordSection1" style="page: WordSection1; caret-color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; word-spacing: 0px; -webkit-text-stroke-width: 0px; text-decoration: none;"><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Hi-<span class="Apple-converted-space"> </span><o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><b class="">Where do I download the PolyPhen_SIFT.db file? I couldn’t create it either because there are problems connecting with the ensemble mysql db<o:p class=""></o:p></b></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">DBI connect('host=<a href="http://ensembldb.ensembl.org/" style="color: blue; text-decoration: underline;" class="">ensembldb.ensembl.org</a>;port=3306','anonymous',...) failed: Unknown MySQL server host '<a href="http://ensembldb.ensembl.org/" style="color: blue; text-decoration: underline;" class="">ensembldb.ensembl.org</a>' (11) at /home/margaret/Projects/ensembl-vep/Bio/EnsEMBL/Registry.pm line 1770.<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">-------------------- EXCEPTION --------------------<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">MSG: Cannot connect to the Ensembl MySQL server at<span class="Apple-converted-space"> </span><a href="http://ensembldb.ensembl.org:3306/" style="color: blue; text-decoration: underline;" class="">ensembldb.ensembl.org:3306</a>; check your settings & DBI error message: Unknown MySQL server host '<a href="http://ensembldb.ensembl.org/" style="color: blue; text-decoration: underline;" class="">ensembldb.ensembl.org</a>' (11)<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><b class="">Here is documentation from the PolyPhen_SIFT plugin file.<o:p class=""></o:p></b></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">--------------------------------------------------------------------------------------------------------------------------<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">A VEP plugin that retrieves PolyPhen and SIFT predictions from a<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">locally constructed sqlite database. It can be used when your main<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">source of VEP transcript annotation (e.g. a GFF file or GFF-based cache)<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">does not contain these predictions.<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">You must either download or create a sqlite database of the predictions.<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">You may point to the file by adding db=[file] as a parameter:<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">--plugin PolyPhen_SIFT,db=[file]<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><span style="background-color: yellow; background-position: initial initial; background-repeat: initial initial;" class="">Human predictions (assembly-independent) are available here:<o:p class=""></o:p></span></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><span style="background-color: yellow; background-position: initial initial; background-repeat: initial initial;" class=""><o:p class=""> </o:p></span></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><span style="background-color: yellow; background-position: initial initial; background-repeat: initial initial;" class=""><a href="https://dl.dropboxusercontent.com/u/12936195/homo_sapiens.PolyPhen_SIFT.db" style="color: blue; text-decoration: underline;" class="">https://dl.dropboxusercontent.com/u/12936195/homo_sapiens.PolyPhen_SIFT.db</a></span> <span class="Apple-converted-space"> </span><b class=""><span style="font-family: Wingdings;" class="">ß</span></b><b class=""><span class="Apple-converted-space"> </span>Not available (no longer exists)<o:p class=""></o:p></b></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">(Please note the download location of this file may change)<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Place this file in $HOME/.vep to have the plugin find it automatically.<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">You may change this directory by adding dir=[dir] as a parameter:<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">--plugin PolyPhen_SIFT,dir=[dir]<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">To create the database, you must have an active database connection<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">(i.e. not using --offline) and add create_db=1 as a parameter:<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><span style="background-color: yellow; background-position: initial initial; background-repeat: initial initial;" class="">--plugin PolyPhen_SIFT,create_db=1</span><o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">--------------------------------------------------------------------------------------------------------------------------<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Best,<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Margaret<o:p class=""></o:p></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div class=""><div style="border-style: solid none none; border-top-width: 1pt; border-top-color: rgb(225, 225, 225); padding: 3pt 0in 0in;" class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><b class="">From:</b><span class="Apple-converted-space"> </span>Dev <<a href="mailto:dev-bounces@ensembl.org" style="color: blue; text-decoration: underline;" class="">dev-bounces@ensembl.org</a>><span class="Apple-converted-space"> </span><b class="">On Behalf Of<span class="Apple-converted-space"> </span></b>Andrew Parton<br class=""><b class="">Sent:</b><span class="Apple-converted-space"> </span>Tuesday, March 10, 2020 3:41 AM<br class=""><b class="">To:</b><span class="Apple-converted-space"> </span>Ensembl developers list <<a href="mailto:dev@ensembl.org" style="color: blue; text-decoration: underline;" class="">dev@ensembl.org</a>><br class=""><b class="">Subject:</b><span class="Apple-converted-space"> </span>Re: [ensembl-dev] Annotations DBs<o:p class=""></o:p></div></div></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div align="center" class=""><table class="MsoNormalTable" border="1" cellspacing="0" cellpadding="0" width="200" style="width: 150pt; background-color: yellow; background-position: initial initial; background-repeat: initial initial;"><tbody class=""><tr class=""><td nowrap="" style="padding: 0.75pt;" class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">USE CAUTION: External Message.<o:p class=""></o:p></div></td></tr></tbody></table></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Hi Margaret,<span class="Apple-converted-space"> </span><o:p class=""></o:p></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">VEP can provide annotations from three sources, which can be specified with a flag on VEP input:<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div><div class=""><ul type="disc" style="margin-bottom: 0in;" class=""><li class="MsoNormal" style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;">—cache - Using a cache is the fastest and most efficient way to use VEP. With a cache file, the vast majority of annotations can be read from this file, reducing the number of required network connections. Cache files can be downloaded using the VEP installer, or can be downloaded manually if required. More information can be found on VEP caches here: <a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ensembl.org_info_docs_tools_vep_script_vep-5Fcache.html-23cache&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=EuPq2OOXcTsL3ZaeT4NARRbtdvaGdc7t2pxc_bsPKDI&e=" style="color: blue; text-decoration: underline;" class="">https://www.ensembl.org/info/docs/tools/vep/script/vep_cache.html#cache</a><o:p class=""></o:p></li><li class="MsoNormal" style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;">—database - VEP can use remote or local database servers to retrieve annotations. <span style="font-family: Helvetica, sans-serif; background-color: white;" class="">By default, VEP is configured to connect to the public Ensembl MySQL instance at<span class="Apple-converted-space"> </span><a href="https://urldefense.proofpoint.com/v2/url?u=http-3A__ensembldb.ensembl.org&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=xxCjIHxXBnPLClgce83JrsJwTZQ2fnTClvIVM513DlA&e=" style="color: blue; text-decoration: underline;" class="">ensembldb.ensembl.org</a>. You can find more information on how to access our database servers, or to set up a local database here: </span><a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ensembl.org_info_docs_tools_vep_script_vep-5Fcache.html-23database&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=oHRqPFmEhohg1l7n6y2vmCf0U3GCfx_Lwl3bNUq207A&e=" style="color: blue; text-decoration: underline;" class="">https://www.ensembl.org/info/docs/tools/vep/script/vep_cache.html#database</a><o:p class=""></o:p></li><li class="MsoNormal" style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;">—custom - If you have annotation data in a standard format such as GFF, VCF or BED, you can integrate this data into your VEP output using our custom annotation functionality. For more information on custom annotations, please see <a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ensembl.org_info_docs_tools_vep_script_vep-5Fcustom.html&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=X15ABtkH0nI3mX9cr5MqzdVf8EcP4xdYXW9bhdHS1c4&e=" style="color: blue; text-decoration: underline;" class="">https://www.ensembl.org/info/docs/tools/vep/script/vep_custom.html</a><o:p class=""></o:p></li></ul><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Let us know if there’s anything else we can do to help.<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Kind Regards,<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Andrew<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div><div class=""><blockquote style="margin-top: 5pt; margin-bottom: 5pt;" class=""><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">On 10 Mar 2020, at 00:40, Linan, Margaret <<a href="mailto:margaret.linan@mssm.edu" style="color: blue; text-decoration: underline;" class="">margaret.linan@mssm.edu</a>> wrote:<o:p class=""></o:p></div></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div><div class=""><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Hi –<span class="apple-converted-space"> </span><o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""> <o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">How do you get the annotations dbs for VEP? I know I can specify them on the command line but they did not come with my installation.<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""> <o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Thanks,<o:p class=""></o:p></div></div><div class=""><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class="">Margaret<o:p class=""></o:p></div></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class="">_______________________________________________<br class="">Dev mailing list </span><a href="mailto:Dev@ensembl.org" style="color: blue; text-decoration: underline;" class=""><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class="">Dev@ensembl.org</span></a><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class=""><br class="">Posting guidelines and subscribe/unsubscribe info:<span class="apple-converted-space"> </span></span><a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__lists.ensembl.org_mailman_listinfo_dev-5Fensembl.org&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=Gixv5gNFUhYwJWf8fi2vtDCO0FkwOwHt21RUS7Xboz4&e=" style="color: blue; text-decoration: underline;" class=""><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class="">https://lists.ensembl.org/mailman/listinfo/dev_ensembl.org</span></a><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class=""><br class="">Ensembl Blog:<span class="apple-converted-space"> </span></span><a href="https://urldefense.proofpoint.com/v2/url?u=http-3A__www.ensembl.info_&d=DwMFaQ&c=shNJtf5dKgNcPZ6Yh64b-A&r=kRxZpbitOhDkEC3BuUN1vDtzo3iicYrRn6woDJL_jnA&m=ckmCwWv2aOHlgURCqjAEFG3NRLLkGe81hdi6gjfQGrU&s=H2c-haOGfvZy_HPHnF7mzI3PUgHmPyg4NUZ3H3FZ0T4&e=" style="color: blue; text-decoration: underline;" class=""><span style="font-size: 9pt; font-family: Helvetica, sans-serif;" class="">http://www.ensembl.info/</span></a><o:p class=""></o:p></div></div></blockquote></div><div style="margin: 0in 0in 0.0001pt; font-size: 11pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></div></div></div></div><span style="caret-color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; word-spacing: 0px; -webkit-text-stroke-width: 0px; text-decoration: none; float: none; display: inline !important;" class="">_______________________________________________</span><br style="caret-color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; text-align: start; text-indent: 0px; text-transform: none; 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