Biology 107 Phage genome annotation progression Lesson 1 Learn about your phage and compare its potential genes with similar phage In this drill you will simply get to know a little bit about the phage whose genome you will be annotating as well as getting some exposure to some of the tools you will use to begin the annotation process Some of the steps you do in this drill today you will probably do frequently throughout the semester as you carefully inspect each gene call in the section of the genome for which you are responsible Get to know your phage Look up your phage by name on the Phages data base PhagesDB page and read the information the discovery student wrote https phagesdb org Bookmark this page Some useful things you could write in your notebook this week What species of bacteria does your phage infect Isolation Host When and where was it found Find a phage with a similar genome sequence On the PhagesDB page use the green Locally Blast this genome Select the BLAST button and wait for the data to show up Your phage is the black line and also the top red line the other red lines are other phages similar to your phage but with different alignment scores Click on any red line or scroll down to the list of phage below ordered by alignment score The higher the score the closer the alignment Click on any score to see the actual nucleotide alignment between your phage and another be sure to note which phages are most closely aligned to your phage that will be useful later See where prediction programs think there are protein coding genes in your phage genome Sign in to PECAAN https discover kbrinsgd org Bookmark this page In the Summary tab start typing in your phage name then select your phage with the associated lab time when the name appears Select the Genes tab at the top Select the Host Trained GeneMark button GeneMark predicts coding potential the likelihood of a stretch of DNA sequence could be encoding a protein A GeneMark document will appear and you can scroll horizontally to look along your entire phage genome Note that there are actually 6 lines the top three are labeled Direct Sequence the bottom three are labeled Complementary Sequence each line represents each coding frame of the DNA 3 forward frames 3 reverse frames Compare the synteny localization of predicted genes of your phage genome sequence to a closely related phage Go back to PECAAN and select the Pham Maps tab This shows the locations of predicted genes colored boxes with numbers and possible tRNAs signs Click and drag or use arrow buttons to look along the genome What do the numbers mean The numbers in the boxes just represent the order in which each gene in that genome appears starting at the front end of the genome Adding a gene or removing a gene will change the numbering of all of the other genes after it The numbers along the black hashed line in the middle represent the nucleotide numbers in kilobases where the 1 represents 1000 nucleotides bases from the front end of the genome What do the colors mean If your gene calls are similar in sequence to genes found in other phages then your gene is considered to be a member of that group of genes called a pham It means that some version of this gene is known to exist to some extent in other phages If you see a gene box that is white it means that the predicted gene is unique it has not been found before in another phage called an orpham What if all of my boxes are all white with no colors are all my genes orphams No sometimes there is a delay between the time a phage genome is sequenced and when it is run through a program called Phamerator which assigns suspected genes to phams PECAAN is updated regularly but a newly sequenced genome like some of ours will not have gene calls assigned to phams right away Eventually they will be so at some point in the semester you should see some if not all of the genes in your phage get assigned to a pham and become a new color You can compare the gene calls to the known genes of other phages that are closely related by using the dropdown menu to select another phage try selecting phage to which your phage was closely related in the BLAST search on the PhagesDB from above when you checked the BLAST search for similar phages a This shows you the alignment of your predicted genes with the genes of another phage Do you see any large gaps There might be a gene in there that wasn t called and should be added Do you see overlaps of genes This may mean a gene was called that should not be there or the start sight is in the wrong place
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