Help:2017 DNA Distribution

Revision as of 17:55, 20 April 2017 by Vinoo (Talk | contribs)


The iGEM 2017 DNA Distribution contains over 1200 high-quality part samples as dried (miniprepped plasmid) DNA. Each sample is QC tested through sequencing and AB test plates.

While there is not enough DNA for 3A Assembly right out of the kit, you can transform a part into competent cells and make your own glycerol stock for future use.

When you receive your Distribution Kit, let us know through twitter (#WelcomeToiGEM). If you're new to the Registry, iGEM, or synthetic biology, you will also want to read through our Help pages before you get started.

Otherwise, read on!


What's included in the 2017 Distribution

Your 2017 DNA Distribution contains the following:
Green BulletPt.png DNA Distribution Kit Plates 1 - 7
Green BulletPt.png Linearized Plasmid Backbones
Green BulletPt.png Competent Cell Test Kit
Green BulletPt.png InterLab Measurement Kit
Green BulletPt.png 2017 Part Submission Kit
Green BulletPt.png iGEM Stickers and Pins!
Green BulletPt.png Sponsor Material

If there is an issue with your distribution kit, please send an email to hq (at) igem . org


DNA Distribution Kit Plates

Quality
In making this year’s distribution, we selected only parts with samples that were sequence confirmed or ends confirmed (long parts > 1600bp). All parts underwent an additional round of our quality control process: sequencing and antibiotic testing. This has ensured a high quality distribution, in which all parts have a sequence result of Confirmed, Partially Confirmed or Long Part. However, be sure to take a look at the quality control results before using a part! Note: We're still in the process of curating the Distribution, and removing samples that did not meet our requirements!

Storage: The distribution kit plates are comprised of dried DNA with cresol red dye, so they are stable at room temperature. However, once the DNA is resuspended in any of the wells, we recommend either storing the kit plate with its plastic cover in a -20C freezer, or aspirating the rest of the resuspended DNA from the well and keeping it separately in a -20C freezer.

Kit Plates 1 - 3

Standards
In addition to the new parts that have been submitted to the Registry, we have transferred over many of our existing sequence verified parts into pSB1C3, the Registry’s standard shipping plasmid backbone. All parts in DNA Distribution Kit Plates 1 to 3 are in pSB1C3.

Why is this useful?

  • all parts are flanked by the BioBrick prefix and suffix.
  • pSB1C3 has our standard primer sites (VF2 and VR), so you can sequence all parts with the same primer set.
  • pSB1C3 has a high copy origin, improving yields for minipreps
  • pSB1C3 is chloramphenicol resistant, so growing and maintaining parts only requires a single antibiotic

Kit Plate 4

DNA Distribution Kit Plate 4 contains parts in a variety of Registry standard plasmid backbones (pSB1C3, pSB1A3, pSB3K3, etc). On this plate you'll find...

  • Registry standard backbones paired with BBa_J04450.
  • Selected parts from 2013 Submissions.
  • Selected parts featured on the Registry

Kit Plate 5

DNA Distribution Kit Plate 5 contains selected parts from 2014 submissions, all in pSB1C3. These selected parts were sequence confirmed or ends confirmed. Please note that, in 2014 we received over 2000 samples, so while Kit Plate 5 only contains a selection from this number, you can request parts not in the distribution during the year.

Kit Plate 6

DNA Distribution Kit Plate 6 contains selected parts synthesized by Gen9, all in pSB1C3. This plate also includes a small selection of PhytoBrick parts.


Competent Cell Test Kit

Confidence
Whether you’re purchasing competent cells or making your own in the lab, you should test their efficiency before you start transforming parts in the Distribution Kit. The Competent Cell Test Kit is a standardized way to test and calculate the efficiency of your competent cells. If you have issues with using the parts in the Distribution Kit, the first thing we’ll need to know is the efficiency of your competent cells.

See the Competent Cell Test Kit page for more in-depth instructions.
Storage: The Competent Cell Test Kit should be stored at -20C, for longer term storage.

Linearized Plasmid Backbones

The 2017 Distribution Kit includes a set of four linearized plasmid backbones: pSB1A3, pSB1C3, pSB1T3, and pSB1K3.m1. These plasmid backbones have been prepared via PCR and purified. Prior to ligation the linearized backbones will need to be digested with with EcoRI, PstI and DpnI (DpnI is optional: to cut up original template DNA used to create linearized plasmid backbone), leaving two ends ready to be ligated to a BioBrick part. All 2017 submissions will need to be in pSB1C3, so we recommend using our pSB1C3 linearized backbone for that purpose.

See the Linearized Plasmid Backbone protocol page for more in-depth instructions on how to use them and make your own.
Storage: The Linearized Plasmid Backbones (pSB1C3/pSB1A3/pSB1T3/pSB1K3: 25ng/ul at 50ul) should be stored at -20C, for longer term storage

InterLab Measurement Kit

New for 2017, we are providing teams with the InterLab Measurement Kit! The InterLab activity is completely optional and is not required for any team. The kit includes 7 tubes (5 with plasmid DNA and 3 with standard reagents):

See the InterLab Measurement Kit page for more in-depth instructions.
Storage: The InterLab_Measurement_Kit should be stored at room temperature or higher 10C. Do not store below 10C as this causes an IRREVERSIBLE precipitation of the LUDOX solution (ex. 4C, -20C)

2017 Submission Kit

For 2017, we are standardizing the part submission shipment: all part submissions must be shipped as dried DNA, in 96 well plate format, and covered with adhesive foil. We've provided a submission kit with all of the materials you'll need to ship your parts.

The Submission Kit includes:

  • (3) 96-well skirted PCR plates
  • (3) universal plate lids
  • (5) adhesive foils
  • bubble wrap
  • a box with the iGEM HQ address, for shipping your submission

See the 2016 Submission Kit for more in-depth instructions.
Storage: The 2017 Submission Kit should be stored at room temperature

Getting Started

Locating a Part in the Distribution

Before using the DNA plates, you should search the Registry for useful parts, which will also tell you if we have sample in stock, its location (if they're in your 2017 Distribution), requirements, quality control, etc.


You can browse the contents of the 2017 Distribution in their entirety by visiting the Repository.

Searching

  • The Catalog of Parts and Devices will allow you to browse parts and devices by various criteria, including function, chassis, standard, etc.
  • The Search Menu will let you search by text or part name.
  • It is also possible to see the contents of the Kit Plates in their entirety. You can look at the DNA Repositories section on the main page.
    • Click on 2017 Distribution to see the contents of all of your kit plates.
    • Click on the 2017 Kit Plate of your choice, which will list all parts by their part name (BBa_..) in a plate along with their quality control information. Or you can click on the small part diagram below each Kit Plate link: "See a summary of the parts in this plate."
    • These options will show you what is in each well of your plate, however they are not the best way to find specific parts you would like to use.

In stock

If you find a part that you would like to use, you need to make sure that a sample of the part is in stock. There are many parts on the Registry that people are still working on, or have decided not to continue working on anymore. This of course means that the DNA is not available in the Registry's Repository. The simplest way to tell whether the part has a sample is to look at the top right of the part's Main Page. If the part has an available sample the top part of the box will be green and say "Sample In stock."

Requesting a part

We've taken care to create a very well-rounded DNA Distribution this spring, however, should you find the part, or parts, that you require are not available in the 2017 DNA Distribution but are available elsewhere in the Registry's Repository, send us an email (hq [AT] igem [DOT] org) in order to request a sample. Include the part name, the plasmid it's located in, the source, and the quality control information, and we'll send it out to you. For more information please see the Requesting Parts page

Using the online QC resources

Here at Registry, we want everyone to take a look at the results of the quality control measures we've taken this year and previous years, in order to make an informed decision when choosing to use a part. We've made sure to update the online repository for the Spring 2017 distribution with our quality control results.

The best way to use our quality control information is to use it on a part by part basis. As you design your project, make sure to check every part that you're interested in for its QC data. After searching for a part in the registry and arriving at its main page, click on the Get This Part link which will take you to the section listing various quality control information for a samples of that part and its location within the registry.

Using the Spring 2017 DNA Distribution

Storage

  • DNA Distribution Kit Plates: The distribution kit plates are comprised of dried DNA, so they are stable at room temperature. However, once the DNA is resuspended in any of the wells, we recommend either storing the kit plate with its plastic cover in a -20C freezer, or aspirating the rest of the resuspended DNA from the well and keeping it separately in a -20C freezer.
  • Competent Cell Test Kit: The Competent Cell Test Kit should be stored at 4C or -20C, for longer term storage.
  • Linearized Plasmid Backbones: The Linearized Plasmid Backbones (pSB1C3/pSB1A3/pSB1T3/pSB1K3: 25ng/ul at 50ul) should be stored at 4C or -20C, for longer term storage.
  • InterLab Measurement Kit: The InterLab Measurement Kit should be stored at room temperature or 4C. Do not store below 4C (e.g. -20C)


DNA Kit Plate Orientation

Locating Your Part from iGEM Videos.

Top view of plates containing dry DNA; red circle indicates well 13H

The foil covers on the (384 well) Kit Plates can be easily punched through with a pipette tip. Unfortunately, the foil cover will also obscure both column and row markings. You can still find your part by correctly orienting the plate using the two notched corners as markers: well A1 is located at the upper left corner of the plate when the long side of the plate with the notched corners is considered the bottom.

Make sure that the two notched corners of the plate are oriented at the BOTTOM of the plate (see the "top view" image on the right for correct orientation)

DNA Kit Plate Instructions

Before you use the DNA in the Distribution Kit Plates, be sure to test the efficiency of your competent cells with the Competent Cell Test Kit.

To use the DNA in the Distribution Kit, follow these instructions:
Note: There is an estimated 2-3ng of DNA in each well, following this protocol, assume that you are transforming with 200-300pg/ul

  1. With a pipette tip, punch a hole through the foil cover into the corresponding well of the part that you want. Make sure you have properly oriented the plate. Do not remove the foil cover, as it could lead to cross contamination between the wells.
  2. Pipette 10uL of dH2O (distilled water) into the well. Pipette up and down a few times and let sit for 5 minutes to make sure the dried DNA is fully resuspended. The resuspension will be red, as the dried DNA has cresol red dye. We recommend that you do not use TE to resuspend the dried DNA.
  3. Transform 1ul of the resuspended DNA into your desired competent cells, plate your transformation with the appropriate antibiotic* and grow overnight.
  4. Pick a single colony and inoculate broth (again, with the correct antibiotic) and grow for 16 hours.
  5. Use the resulting culture to [http://openwetware.org/wiki/Miniprep/Kit-free_high-throughput_protocol miniprep] the DNA AND make your own glycerol stock (for further instruction on making a glycerol see [http://openwetware.org/wiki/Endy:Making_a_long_term_stock_of_bacteria this page]). We recommend using the miniprepped DNA to run QC tests, such as restriction digests and sequencing.

* To know which antibiotics to use, look at the plasmid that the part is in. The naming scheme for plasmids is specifically designed to indicate antibiotic resistance.

Note: There is not enough DNA in each well to perform anything but transformations


Get & Use...

Register | Locating Parts | Quality Control | Distribution Kit | Requesting Parts | Using Parts