{"action":"created","issue":{"url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/7","repository_url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization","labels_url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/7/labels{/name}","comments_url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/7/comments","events_url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/7/events","html_url":"https://github.com/ohsu-computational-biology/tcrseq_normalization/issues/7","id":152611982,"number":7,"title":"Determine Amplification Bias - ei","user":{"login":"weshorton","id":16656587,"avatar_url":"https://avatars.githubusercontent.com/u/16656587?v=3","gravatar_id":"","url":"https://api.github.com/users/weshorton","html_url":"https://github.com/weshorton","followers_url":"https://api.github.com/users/weshorton/followers","following_url":"https://api.github.com/users/weshorton/following{/other_user}","gists_url":"https://api.github.com/users/weshorton/gists{/gist_id}","starred_url":"https://api.github.com/users/weshorton/starred{/owner}{/repo}","subscriptions_url":"https://api.github.com/users/weshorton/subscriptions","organizations_url":"https://api.github.com/users/weshorton/orgs","repos_url":"https://api.github.com/users/weshorton/repos","events_url":"https://api.github.com/users/weshorton/events{/privacy}","received_events_url":"https://api.github.com/users/weshorton/received_events","type":"User","site_admin":false},"labels":[{"url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/labels/bug","name":"bug","color":"ee0701"}],"state":"open","locked":false,"assignee":null,"assignees":[],"milestone":null,"comments":1,"created_at":"2016-05-02T18:23:05Z","updated_at":"2016-08-02T21:14:26Z","closed_at":null,"body":"### Summary\r\nIn PCR1, we use a multiplex reaction of 20 forward and 13 reverse primers. Each primer has a different amplification rate during the reaction, and we need to characterize each amplification rate.\r\n+ Example\r\n     + Clonotype A is amplified by V1J1 primer combination, Clonotype B is amplified by V2J2 combination.\r\n     + Sample contains equal starting amounts of Clonotype A and Clonotype B\r\n     + Post-amplification/sequencing, sample contains twice as many counts of Clonotype A than Clonotype B\r\n     + V1J1 has double the efficiency of V2J2\r\n\r\n### Significance\r\nWe can't know the initial counts of clonotypes within a sample, and can only measure them post-amplification and sequencing. We need to incorporate this into our model ([Wiki - Model](https://github.com/ohsu-computational-biology/tcrseq_normalization/wiki)) in order to determine the correct count for each clonotype.\r\n\r\n### To Do\r\n1. Determine primer independence (link to issue)\r\n2. Depending on above, calculate 260 ei factors, or 33\r\n\r\n### Approach\r\n1. Control Experiment\r\n     + Samples containing:\r\n          + Constant spike concentration (To Do: what is this value?)\r\n          + NO gDNA\r\n     + 20 total samples\r\n\r\n2. Data Analysis and Interpretation\r\n     + Boxplots of spike counts\r\n          + for all 260 primer combinations\r\n          + grouped by V primer (20)\r\n          + grouped by J primer (13)\r\n     + ANOVA?\r\n          + predictor variable - primer (or primer combination)\r\n          + response variable - count\r\n          + Ho: No variation between counts for different primers\r\n          + Ha: Spike count deviates significantly from mean count depending on primer used.\r\n     + Difference from mean used to calculate ei factor\r\n          + How?\r\n\r\n3. Relevant plots, tables, and other information\r\n     + [Box plots] of spike counts vs primer identity\r\n     + [Table] of ANOVA results\r\n     + [Table] of ei factors\r\n"},"comment":{"url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/comments/237046178","html_url":"https://github.com/ohsu-computational-biology/tcrseq_normalization/issues/7#issuecomment-237046178","issue_url":"https://api.github.com/repos/ohsu-computational-biology/tcrseq_normalization/issues/7","id":237046178,"user":{"login":"weshorton","id":16656587,"avatar_url":"https://avatars.githubusercontent.com/u/16656587?v=3","gravatar_id":"","url":"https://api.github.com/users/weshorton","html_url":"https://github.com/weshorton","followers_url":"https://api.github.com/users/weshorton/followers","following_url":"https://api.github.com/users/weshorton/following{/other_user}","gists_url":"https://api.github.com/users/weshorton/gists{/gist_id}","starred_url":"https://api.github.com/users/weshorton/starred{/owner}{/repo}","subscriptions_url":"https://api.github.com/users/weshorton/subscriptions","organizations_url":"https://api.github.com/users/weshorton/orgs","repos_url":"https://api.github.com/users/weshorton/repos","events_url":"https://api.github.com/users/weshorton/events{/privacy}","received_events_url":"https://api.github.com/users/weshorton/received_events","type":"User","site_admin":false},"created_at":"2016-08-02T21:14:26Z","updated_at":"2016-08-02T21:14:26Z","body":"To Do\r\n1. Turn into milestone\r\n2. How is amplification bias captured in our new normalization method?\r\n + Talk with Burcu\r\n3. If possible, determine general amplification bias. If not, must determine for each sample/batch."}}
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