ApE Tutorial Video #4 - Restriction digests

00:07:42
https://www.youtube.com/watch?v=UYy952xwyE0

Ringkasan

TLDRThis video is part of the Ape tutorial series, focusing on how to handle restriction enzyme sites and perform virtual gel electrophoresis. Users can highlight, select, and clear restriction enzyme sites, with functionalities available to simulate enzyme digests. The video guides through processes such as setting up single or double digests, observing the resultant bands on an agarose gel, and interpreting the data such as sizes and positions of the restriction sites. Further features include reordering lanes, copying lanes between different windows, setting up complex gels, and partial digestion simulations. The tutorial also provides instructions on saving the gel diagrams in various formats for documentation purposes.

Takeaways

  • 🔍 How to clear highlighting on enzyme sites.
  • 🔬 Perform restriction enzyme digests and visualize on virtual gel.
  • 🔢 See size and position of bands by hovering over them on the gel.
  • 🔄 Reorder lanes through drag and drop.
  • 📊 Set up complex gels using the digestion dialogue.
  • 🔗 Option to perform single, double, or triple digests.
  • 📂 Save gel diagrams in various formats for further use.
  • 📋 Copy lane information between digest windows.
  • ➗ Simulate partial digests by adjusting digestion percentages.
  • 📃 Access comprehensive information about bands and lanes.
  • 🖨️ Export the virtual gels as EPS, SVG, PDF, or PowerPoint.
  • 💡 Additional setup features for enzyme digests.

Garis waktu

  • 00:00:00 - 00:07:42

    In this fourth video of the APE tutorial series, the focus is on managing and manipulating restriction enzyme sites in sequence files. The video begins by reviewing how to clear highlighting of restriction sites, then introduces performing restriction digests. By selecting enzymes such as Eco R1 and HindIII, users can create a virtual gel to see the potential results, including band sizes and positions on an agarose gel. The video demonstrates the process of single and double digests, explains how to reorder lanes, and describes setting up complex gels using the digestion dialogue. This enables simultaneous setup of multiple lanes and customization of digests, including partial digest options and adding enzyme columns. Once satisfied, users can save or export the digest information in various formats for documentation.

Peta Pikiran

Video Tanya Jawab

  • How do I remove highlighting from the sequence file?

    You can remove highlighting by selecting 'clear highlighting' in the enzymes menu or using the shift-click option on the toolbar button.

  • What can be done with restriction enzymes in this tool?

    You can perform restriction enzyme digests and visualize them on a virtual gel.

  • How can I view the size and base pairs of the bands on the gel?

    Hovering the mouse over the virtual bands shows the size in base pairs along with highlighting other properties.

  • Can I perform both single and double enzyme digests?

    Yes, you can do both single and double digests, and visualize them individually or together.

  • How can I reorder the lanes on the virtual gel?

    You can click and drag to reorder the lanes on the virtual gel.

  • Is it possible to set up more complex gels at once?

    Yes, using the digestion dialogue, you can set up multiple lanes and enzymes for complex gels.

  • How can I save the virtual gel information?

    The gel can be saved as an encapsulated postscript, SVG, PDF, or PowerPoint slide.

  • What format options are available for saving the gel?

    You can save the gel in encapsulated postscript, SVG, PDF, or PowerPoint slide formats.

  • How do I copy lane information to another digest window?

    You can drag a lane from one digest window to another to copy it.

  • Can I visualize partial digests?

    Yes, you can set up partial digests in the digestion dialogue by adjusting the digestion percentage.

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Teks
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Gulir Otomatis:
  • 00:00:00
    welcome to the fourth video in the ape
  • 00:00:02
    video tutorial series
  • 00:00:05
    in the last
  • 00:00:06
    video we
  • 00:00:08
    did highlighting of and selection of
  • 00:00:11
    restriction enzyme sites
  • 00:00:13
    these sites
  • 00:00:15
    we can remove the highlighting by doing
  • 00:00:18
    clear highlighting within the enzymes
  • 00:00:19
    menu or shift click within this toolbar
  • 00:00:22
    button here
  • 00:00:24
    that will remove all of the highlighting
  • 00:00:26
    from that sequence file
  • 00:00:31
    you can
  • 00:00:33
    also do other things with restriction
  • 00:00:35
    enzymes
  • 00:00:36
    for example
  • 00:00:37
    you can do restriction digests
  • 00:00:40
    so if you select
  • 00:00:43
    eco r1 and hindi 3 you can do a digest
  • 00:00:47
    as a double digest and show
  • 00:00:49
    [Music]
  • 00:00:50
    a virtual
  • 00:00:52
    gel
  • 00:00:53
    of what it would look like to digest
  • 00:00:55
    this restriction this sequence with
  • 00:00:58
    those two restriction enzymes
  • 00:01:01
    you see
  • 00:01:03
    virtual
  • 00:01:04
    bands on this agarose gel
  • 00:01:07
    and if you put your mouse over the band
  • 00:01:10
    you can see the size
  • 00:01:12
    and base pairs and you can also see
  • 00:01:17
    red highlighting that shows you the
  • 00:01:21
    size
  • 00:01:22
    the
  • 00:01:24
    left restriction sites the position of
  • 00:01:26
    that restriction sites the right
  • 00:01:29
    restriction site the position of that
  • 00:01:31
    and the mass percent of that band as a
  • 00:01:34
    percent of the total mass
  • 00:01:36
    of the digest
  • 00:01:38
    if you click on the band or on the
  • 00:01:42
    fan text
  • 00:01:43
    it will highlight that band
  • 00:01:45
    within the sequence
  • 00:01:48
    you can also
  • 00:01:50
    find that
  • 00:01:53
    band
  • 00:01:54
    by
  • 00:01:55
    putting your mouse over this small map
  • 00:01:58
    that shows the features
  • 00:02:00
    of the sequence as well as the position
  • 00:02:02
    of each of the restriction enzyme sites
  • 00:02:08
    you can
  • 00:02:09
    rather than doing a double digest you
  • 00:02:11
    can do a digest with all which will
  • 00:02:13
    digest individually
  • 00:02:19
    with eco r1 or with hindi 3 separately
  • 00:02:23
    if you put your mouse now over either of
  • 00:02:26
    these bands
  • 00:02:27
    it will change the text
  • 00:02:29
    to the left side
  • 00:02:31
    to a list of
  • 00:02:33
    restriction
  • 00:02:34
    bands for that digest
  • 00:02:39
    and will update as you highlight
  • 00:02:42
    different bands
  • 00:02:44
    if you click in this show in lane
  • 00:02:46
    information box
  • 00:02:48
    you can get a list of um
  • 00:02:52
    all of the lanes the ladder lane
  • 00:02:55
    here
  • 00:02:56
    lane two which is an eco-r one digest
  • 00:03:00
    and lane three which is a hindi
  • 00:03:03
    digest
  • 00:03:05
    if you
  • 00:03:06
    click and drag
  • 00:03:08
    on any one of these lanes you can
  • 00:03:11
    reorder
  • 00:03:13
    the
  • 00:03:14
    position within the
  • 00:03:16
    of the digests
  • 00:03:18
    within the gel
  • 00:03:20
    if you have multiple restriction digests
  • 00:03:22
    open at once
  • 00:03:28
    for example if i had an eco or one
  • 00:03:30
    digest
  • 00:03:32
    in one window
  • 00:03:34
    and a handy three digest in another
  • 00:03:35
    window
  • 00:03:36
    you can
  • 00:03:39
    drag
  • 00:03:41
    one
  • 00:03:43
    lane from one digest window to a
  • 00:03:45
    different digest window
  • 00:03:48
    and it will copy it from this window to
  • 00:03:50
    this window
  • 00:03:53
    if you'd like to set up more complex
  • 00:03:55
    gels all at once rather than doing an
  • 00:03:58
    individual digest you can use the
  • 00:04:00
    digestion dialogue
  • 00:04:03
    this allows you to set up multiple lanes
  • 00:04:06
    all at once rather than moving them
  • 00:04:08
    around from one window to another
  • 00:04:11
    in this digest in this dialogue each of
  • 00:04:13
    the lanes of the new digest is
  • 00:04:16
    represented by a row
  • 00:04:18
    and the possible restriction enzymes
  • 00:04:21
    are columns
  • 00:04:23
    so in each lane you can choose either to
  • 00:04:26
    have
  • 00:04:27
    nothing
  • 00:04:28
    you can choose to have a ladder from
  • 00:04:30
    your
  • 00:04:32
    predefined ladder set
  • 00:04:34
    those can be edited with it
  • 00:04:38
    from the
  • 00:04:39
    edit from the enzymes menu under the
  • 00:04:43
    ladder editing dialog
  • 00:04:46
    or you can choose to digest
  • 00:04:48
    one of the presently open windows
  • 00:04:52
    so in this case we'll do the first lane
  • 00:04:55
    as a ladder
  • 00:04:57
    the second lane will be an eco r1 digest
  • 00:05:00
    of our plasmid
  • 00:05:02
    you can add additional lanes with the
  • 00:05:04
    plus button here
  • 00:05:07
    and it will make a copy of the lane that
  • 00:05:10
    you just
  • 00:05:11
    were in
  • 00:05:13
    you can
  • 00:05:15
    change from digesting
  • 00:05:18
    with eco r1 to handy 3.
  • 00:05:21
    you can also set
  • 00:05:22
    up potentially
  • 00:05:26
    partial digests if you would like to see
  • 00:05:29
    what a partial digestion would look like
  • 00:05:31
    you can change the percentages from 100
  • 00:05:34
    digestion to 90 or any
  • 00:05:38
    any value of digestion
  • 00:05:42
    you can select all on the column or
  • 00:05:45
    deselect all in the column from here you
  • 00:05:47
    can add additional enzyme
  • 00:05:50
    columns with this button so if you would
  • 00:05:53
    like to make a bam h1 digest
  • 00:05:58
    you can do it like this
  • 00:06:02
    there's a convenience feature for
  • 00:06:05
    digesting any particular dna with each
  • 00:06:07
    of the enzymes
  • 00:06:09
    and it will set up
  • 00:06:12
    from all of the enzymes that you have at
  • 00:06:14
    the
  • 00:06:15
    have a column for it will set up an
  • 00:06:17
    individual digest
  • 00:06:18
    for each of those
  • 00:06:24
    you can do
  • 00:06:26
    double digest or triple digest in any
  • 00:06:28
    particular lane as well
  • 00:06:31
    then when you're satisfied with the lane
  • 00:06:34
    information as you've set it up click ok
  • 00:06:37
    to get a full
  • 00:06:39
    digest pattern
  • 00:06:41
    of each of those and again you can
  • 00:06:43
    highlight to find the size of any of
  • 00:06:45
    these bands
  • 00:06:46
    you can reorder the bands
  • 00:06:50
    you can get a list of all of the
  • 00:06:54
    information
  • 00:06:56
    you can copy that text and paste it into
  • 00:06:59
    your notebook your electronic notebook
  • 00:07:03
    or you can
  • 00:07:12
    you can copy the
  • 00:07:14
    you can save the um file as a
  • 00:07:23
    encapsulated postscript an svg
  • 00:07:25
    a pdf or a powerpoint slide
  • 00:07:28
    and
  • 00:07:30
    get the information in a graphical
  • 00:07:32
    format
  • 00:07:35
    that's all for this video of the
  • 00:07:38
    tutorial video series
Tags
  • restriction enzymes
  • virtual gel
  • digestion
  • highlighting
  • enzyme sites
  • double digest
  • partial digest
  • lane reordering
  • data analysis
  • formats