Uploading trace files to bold
a. As you now know, the sequencing outputs a folder of files. The files you are interested in have an extension .ab1, e.g., LOP001-11_F.ab1. These raw files (traces) can be edited into the form we are use to seeing DNA sequences represented in, i.e., a string of letters. However, as editing can be a subjective task, BOLD also requires the raw files (traces) be uploaded as part of a DNA barcode’s collateral data.
b. To add trace files of your new sequences to the appropriate records in BOLD create a folder on your desktop called Traces and place in it all the .ab1 files that you would like to upload. [Note: you probably already have this folder from earlier, see 9. SANGER SEQUENCE EDITING]
c. To create a list of the files in the Traces folder open a terminal window. On a Windows computer type "cmd" into the search box in the "start" menu and click on the cmd program to open it. To navigate to the Traces folder, type cd desktop, press enter, type cd Traces, press enter. Next type dir > list.txt, press enter. [Note: you already used this trick for the images upload]
d. Download the files from here and save the file as data.xls in the Traces folder on the desktop.
e. Open data.xls in MS Excel.
f. You can then open list.txt and move the data into the Filename (.ab1) column in data.xls. The cells in this column
should contain the name of a trace file including the extension (.ab1 ) (two rows are provided as an example).
g. In the FORWARD PCR PRIMER column enter the registered name of the forward primer used during the PCR. Copy it down through the entire column to the end of your file list.
h. In the REVERSE PCR PRIMER column enter the registered name of the reverse primer used during the PCR. Copy it down through the entire column to the end of your file list.
i. In the column SEQUENCING PRIMER enter the registered name of your sequencing primer. It will alternate between forward and reverse. For example, LCO1490 should line up with the read direction Forward.
j. In the Read Direction column enter Forward or Reverse depending on the direction of the .ab1 files it refers to.
k. You need to check and carefully insert your Process ID's from your BOLD project into the Process ID column. Make sure they match up with the correct specimens. The Process IDs are the codes under Sequence Page in your Project Console on BOLD.
l. The Marker is COI-5P for all samples.
m. Save the file. [Check: file name is data.xls and it is in your folder Traces]
n. Delete list.txt from your Traces folder.
o. The folder Traces needs to be zipped before submission to BOLD. See 5. SPECIMEN IMAGES AND UPLOAD TO BOLD for details on how to do this. Save as Traces.zip.
p. Navigate to your BOLD project’s Project Console and under the Uploads menu on the left click Trace Files . Browse through to Traces.zip and click Submit.
b. To add trace files of your new sequences to the appropriate records in BOLD create a folder on your desktop called Traces and place in it all the .ab1 files that you would like to upload. [Note: you probably already have this folder from earlier, see 9. SANGER SEQUENCE EDITING]
c. To create a list of the files in the Traces folder open a terminal window. On a Windows computer type "cmd" into the search box in the "start" menu and click on the cmd program to open it. To navigate to the Traces folder, type cd desktop, press enter, type cd Traces, press enter. Next type dir > list.txt, press enter. [Note: you already used this trick for the images upload]
d. Download the files from here and save the file as data.xls in the Traces folder on the desktop.
e. Open data.xls in MS Excel.
f. You can then open list.txt and move the data into the Filename (.ab1) column in data.xls. The cells in this column
should contain the name of a trace file including the extension (.ab1 ) (two rows are provided as an example).
g. In the FORWARD PCR PRIMER column enter the registered name of the forward primer used during the PCR. Copy it down through the entire column to the end of your file list.
h. In the REVERSE PCR PRIMER column enter the registered name of the reverse primer used during the PCR. Copy it down through the entire column to the end of your file list.
i. In the column SEQUENCING PRIMER enter the registered name of your sequencing primer. It will alternate between forward and reverse. For example, LCO1490 should line up with the read direction Forward.
j. In the Read Direction column enter Forward or Reverse depending on the direction of the .ab1 files it refers to.
k. You need to check and carefully insert your Process ID's from your BOLD project into the Process ID column. Make sure they match up with the correct specimens. The Process IDs are the codes under Sequence Page in your Project Console on BOLD.
l. The Marker is COI-5P for all samples.
m. Save the file. [Check: file name is data.xls and it is in your folder Traces]
n. Delete list.txt from your Traces folder.
o. The folder Traces needs to be zipped before submission to BOLD. See 5. SPECIMEN IMAGES AND UPLOAD TO BOLD for details on how to do this. Save as Traces.zip.
p. Navigate to your BOLD project’s Project Console and under the Uploads menu on the left click Trace Files . Browse through to Traces.zip and click Submit.
UPLOADING FASTA FILES TO BOLD
a. Open the file sequences.fasta in MS Word. Use right click Open with.
b. You need to replace all the sequence names with the appropriate Sample ID or Process ID. [Note: having the BOLD Project Console open will help with this][Note: for this workshop purpose the Sample ID is of the form (YourInitials)A1]
c. Under the Edit menu, click Select All then under Edit, click Copy.
d. Navigate to your BOLD project’s Project Console and under the Uploads menu on the left click Sequences.
e. Right click on the box PASTE SEQUENCES IN FASTA FORMAT: and click Paste.
f. Select the Markers: as COI-5P and select or register your Run site.
b. You need to replace all the sequence names with the appropriate Sample ID or Process ID. [Note: having the BOLD Project Console open will help with this][Note: for this workshop purpose the Sample ID is of the form (YourInitials)A1]
c. Under the Edit menu, click Select All then under Edit, click Copy.
d. Navigate to your BOLD project’s Project Console and under the Uploads menu on the left click Sequences.
e. Right click on the box PASTE SEQUENCES IN FASTA FORMAT: and click Paste.
f. Select the Markers: as COI-5P and select or register your Run site.