The Commands menu is available by either pressing the Commands button at the top of the contig editor window, or by pressing the Control key and the left mouse button, or by pressing right mouse button with the mouse cursor anywhere within the sequence display section of the contig editor. A menu will be revealed containing the following options (which are described in greater detail below).
This Contig Editor Commands menu function Performs a search. See section Searching.
This Contig Editor Commands menu function Creates an annotation. See section Annotations.
This Contig Editor Commands menu function Edits an annotation. See section Annotations.
This Contig Editor Commands menu function Removes an annotation. See section Annotations.
This Contig Editor Commands menu function writes any edited data to disk. The undo history is cleared and it is no longer possible to quit and abandon these saved changes. The Control-x followed by Control-s will also save the contig editor in the same manner as the Save command.
This Contig Editor Commands menu function outputs the current contig, as currently shown (e.g. with status lines) to a file. The user can select the region to dump, the length of each line, and the file name to use. The sequence names can be up to 40 characters, but often projects do not use the full length. To avoid wasted space in the output the number of columns to use for sequence names can be adjusted.
This Contig Editor Commands menu function produces a trace file for the consensus sequence by averaging the traces of the readings. The command brings up a dialogue containing controls to specify the filename, the consensus start and end positions, the strand, and whether to use matching reads.
As the trace of a reading is dependent on the direction it was read, the consensus trace can be computed from all the reads in either the forward or reverse directions, but not both at once. When the "Use only matching reads" toggle is set to "Yes" only the readings of the correct strand that have the same base call as the consensus sequence are used. The option is useful for producing wild-type trace files for a mutation analysis project.
This Contig Editor Commands menu function operates in a very similar manner to the main Gap4 List Confidence command (see section List Confidence), except that it only operates on the current contig, and it uses the current editor consensus confidences rather than the ones saved to disk. It displays a dialogue requesting a range within the contig and a question asking if only summary of the results is required.
Pressing OK or Apply will add to the editor information line a count of the expected number of errors and the error rate. If the "Only update information line" question was answered "No" then the full frequency table will also be output. It will appear in the main text output window in the same format as the "List Confidence" command in the main Gap4 View menu. The Apply button can be used to calculate the number of errors without removing the dialogue.
It is often the very ends of contigs (which are generally low coverage and bad quality) that have most of the errors, and so it is sometimes useful to set a range which includes all of the contig except for around 1000 bases from each end.
This Contig Editor Commands menu function is used to produce a list of all the bases annotated with mutation tags (or those bases which differ from the consensus/reference sequence). If the tags or differences are within segments of sequence which are also annotated with EMBL feature table CDS records, the report will include data describing its effect. The report, which can be sorted by sequence or position, includes the reading names, mutation positions relative to the reference sequence, the actual change, its effect, and the evidence. An example is shown below.
001321_11aF 33885T>Y (silent F) (strand - only) 001321_11aF 34407G>K (expressed E>[ED]) (strand - only) 001321_11cF 35512T>Y (silent L) (double stranded) 001321_11cF 35813C>Y (expressed P>[PL]) (double stranded) 001321_11dF 36314A>R (expressed E>[EG]) (double stranded) 001321_11eF 36749A>R (expressed K>[KR]) (double stranded) 001321_11eF 37313T>K (noncoding) (strand - only) 000256_11eF 36749A>G (expressed K>R) (double stranded)
Here the first record is for reading 001321_11aF, position 33885, T changed to T and C (i.e. is heterozygous) to produce no amino acid change, with evidence coming only from the complementary strand. The last record is for reading 000256_11eF, position 36749, A changed to G, producing an amino acid change K to R, with evidence from both strands of the sequence. The penultimate record denotes a heterozygote in a noncoding region.
This Contig Editor Commands menu function allows the user to employ the primer selection algorithm OSP to find primers for sequencing experiments. See section Primer Selection.
This Contig Editor Commands menu function performs a sequence alignment between the currently selected segment of a reading and the consensus sequence. It provides a simple way of extending the visible part of a reading to use its hidden data, which is often useful to double strand a short section of consensus without the need to perform further experiments. On a sequence, highlight the cutoff data to align along with a small section of the good quality non-cutoff data. Then select the align command and adjust the cutoff point as desired. Pads are inserted in the consensus and readings as necessary, although pads will not be inserted in the cutoff data of other sequences.
One weakness of the alignment strategy used in the current shotgun assembly algorithm is that padding characters are not always aligned with one another. This Contig Editor Commands menu function attempts to align padding characters using a very simply strategy. It does not solve all pad alignment problems but is a useful first step during cleaning-up operations. After shuffling, all columns consisting entirely of pads are removed.
This Contig Editor Commands menu function marks a reading for subsequent removal. See section Removing Readings
This Contig Editor Commands menu function breaks the contig so that the reading underneath the editing cursor is the left end of a new contig. In order to perform this operation all edits are saved automatically first. Once saved these edits cannot be undone. This operation is identical to the Break Contig command in the main menu.See section Break Contig.