Difference between revisions of "Help:2021 DNA Distribution"

Latest revision as of 21:39, 8 July 2021

	The 2021 DNA Distribution Kit
	Whether you’re new to iGEM and the Registry of Standard Biological Parts or an experienced participant, make sure to read through the Distribution Handbook.

The iGEM 2021 DNA Distribution contains over 2000 high-quality part samples as dried plasmid DNA. Each sample is QC tested through sequencing and AB test plates. While there is not enough DNA for 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.

Lastly, for 2022 and beyond we will be making a second-generation iGEM Distribution, designed anew and completely rebuilt using synthesis! We are asking you, our iGEM teams and community, to tell us what you think about the existing or previous Distributions, and what you wish to be true about the iGEM Distribution 2.0.

What's included in the 2021 Distribution

Your 2021 DNA Distribution contains the following:
DNA Distribution Kit Plates 1 - 6
iGEM Stickers and Pins!
iGEM Competition Information!
Sponsor Material

Note: Unfortunately, due to the COVID-19 pandemic and limitations to lab access, we were not able to prepare some kit components. However, we have provided resources on how you can make/obtain your own. We apologize for any inconvenience.

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

DNA Distribution Kit Plates

The distribution only contains parts with samples that are sequence confirmed or ends confirmed (long parts > 1600bp). This has ensured a high quality distribution, in which all parts have a sequence result of Confirmed, Partially Confirmed/Single Error or Long Part. All users should review the sequence results of a part sample before using it.

The majority of part samples in the in the Distribution Kit are in pSB1C3, the Registry’s standard shipping plasmid backbone.

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

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 -20°C freezer, or aspirating the rest of the resuspended DNA from the well and keeping it separately in a -20°C freezer.

Note: The distribution kit plates for 2021 (labeled as v2.19) are a replication of the 2019 source plates. We will be reuploading sequencing data for the 2021 plates shortly, but for now, please reference the corresponding 2019 kit plates.

Additional Resources

Due to the COVID-19 pandemic and limitations to lab access, we were not able to prepare some distribution kit components. The resources below offer guidance on how you can make/obtain your own.

Competent Cell Efficiency

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. If you've purchased your competent cells then they will likely come with a 10 pg/ul control. If you do not have a readily available control, please see the Competent Cell Test Kit page on how to make your own and calculate the efficiency of your competent cells.
If you have issues with assembly and cloning, the first thing we’ll need to know is the efficiency of your competent cells.

Linearized Plasmid Backbones

Linearized plasmid backbones are prepared via PCR and purified as an easy way to do assembly/cloning. Prior to ligation the linearized backbones will need to be digested 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.
See the Linearized Plasmid Backbone protocol page for more in-depth instructions on how make your own and use them.

Measurement Kit

The iGEM Engineering Committee] has put together resources on the protocols and components you need for the calibration of plate readers for fluorescent intensity and cell density measurements if you are measuring fluorescence when testing your devices.

Sponsor Offers

2021 iGEM Teams also have access to sponsor offers for synthesis from IDT, TWIST Bioscience, and GenScript. We highly encourage our teams to use these offers, from creating a library of parts to synthesizing their constructs/devices entirely.

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 2021 Distribution), requirements, quality control, etc.

You can browse the contents of the 2021 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 2021 Distribution to see the contents of all of your kit plates.
  • Click on the 2021 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."

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 2021 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 samples of that part and their locations within the Registry.

Using the Spring 2021 DNA Distribution

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. When following this protocol, assume that you are transforming with 200-300pg/µL

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 10µL 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 1µL 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...