MGH   
CCIB
 
Complete Plasmid Sequencing: Sample Submission

PLEASE REVIEW OUR GUIDELINES BEFORE SUBMITTING YOUR PLASMID(S) !

To ensure the successful sequencing and assembly of your plasmid(s), all guidelines for sample preparation and submission must be addressed. Please contact us if these requirements cannot be met so we can work with you on identifying a solution on a case-by-case basis.

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Sample Requirements:

  • Please submit CIRCULAR DNA only! Linear DNA (such as amplicons and restriction fragments) have to be submitted for our Complete Amplicon Sequencing service, as linear DNA samples require a slightly modified assembly approach.

  • To ensure suitable sample quality, the use of a column-based purification method is preferred.

  • Contamination with bacterial genomic DNA must be avoided as it is the most common reason for assembly failure.

  • The purified plasmid DNA should be resuspended in nuclease-free water, in 10 mM Tris-HCl (pH 7.5-8.5), or in LTE Buffer (10 mM Tris-HCl, pH 8.0, 0.1 mM EDTA).

  • The plasmid DNA isolate must be diluted to a final concentration of 40-65 ng/µl.
    Please note: The DNA concentration should be determined by spectrophotometry. Our recommendations for accurately determining DNA concentration can be found here.

  • Our automated workflow requires the submission of 35 µl of the diluted plasmid DNA.


Tube Orders:

  • If you are submitting your samples in tube format, please use standard 1.5 ml flip-cap Eppendorf-type tubes.
    Please note: Our operational setup does not accept 0.2 ml tubes, 0.5 ml tubes, strip-tubes, and screw-cap tubes.

  • Please label the top of the lid of each tube - as clearly as possible - with a maximum of 5 alpha-numeric characters (for example: 5B001). We strongly recommend to hand-write each label using a permanent marker. Self-adherent stickers might fall off during shipment, thereby making sample identification impossible.

  • We recommend wrapping tubes with Parafilm to prevent samples from drying.

  • Tubes should be submitted in order as listed on the submission form (not randomly placed, and not loose/unracked).
    Please note: If you are using one of our remote drop-off locations, please rack your tubes in one of the provided sample boxes and attach order form with a rubber band.
    If for some reason no boxes should be available, please tape tubes to the order form (or a separately attached sheet) in the same order as listed on your order form. You may also use any other type of rack/box designed to hold 1.5 ml tubes.

  • If your order contains 48-95 samples or is a full 96 sample plate, please use the plate format to receive a discount (see plate guidelines below).

 

Plate Orders. These guidelines apply to both the 48-95 and full 96 sample plate discounted rates

  • If you are submitting your samples in plate format, please use only V-bottom PCR plates.

  • We recommend the use of semi-skirted (or skirted) plates to prevent the PCR plate from bending during transit, which could result in loosening of the seal and subsequent sample loss and cross-contamination.

  • Please arrange your samples horizontally (in rows) in the PCR plate. Proceed from well A1 to A12, from B1 to B12, from C1 to C12, etc.

  • We accept partial plates but please do not leave any empty wells between samples, and please do not leave any empty spaces between samples on the order form.

  • No discounted rate will be applied for orders with fewer than 48 samples, even when submitted in plate format. Multiple orders cannot be combined to receive either the 48-95 or the full 96 plate discount.

  • The order form must match the order and position of samples in the PCR plate.

  • Please label the plate as clearly as possible - with a maximum of 5 alpha-numeric characters (for example: 5B001).

  • Tightly seal your plate(s) with an adhesive sheet/foil to prevent samples from drying and/or cross-contamination.

 

Sample Shipment:

We are currently offering our Complete Plasmid Sequencing service for individual samples and full plates once a week, beginning each Wednesday. The deadline for sample arrival at the Core Facility (38 Sidney St. Cambridge) is 4pm. If you should miss this cutoff time, your samples will be entered into the queue and processed during the following cycle.

Samples may be submitted early and they will be stored at the core under the appropriate conditions until the Wednesday start date. Full plates will be processed on a first come, first serve basis and the turnaround time will depend on the current demand. We ask researchers to notify the NGS Team Manager ahead of time to allow us to plan accordingly.

Please submit your sample(s), including a hard copy of the completed order form, by using one of our remote dropboxes (please see here) or by delivering it personally to our core facility.

If remote or direct sample drop-off should not be an option for you, please ship your samples, including a hard copy of the completed order form, via a courier delivery service of your choice (for example, overnight shipment via FEDEX, UPS, etc.) to our core facility. Samples can be shipped at ambient temperature.

Shipping Tubes:

Unfortunately, we often receive sample tubes that were damaged during shipment (cracked or shattered tube, sample loss due to leakage or evaporation). Therefore, we strongly recommend sealing the sample tube with Parafilm and further protecting it with bubble wrap or several layers of Kimwipe tissue. Another option is to place the sample tube into a secondary container such as a Falcon tube or small cardboard or plastic box.

Shipping 96-well Plates:

Samples should be arrayed in a 96-well (semi-skirted or skirted) plate. To prevent sample loss and/or cross contamination, we recommend tightly sealing all wells of the plate with Microtube Caps. PCR Tube Strip Flat Caps (Eppendorf cat# 0030124847 or VWR cat# 75874-710) work well for most plates. Our everyday experience tells us that plate foils/plastic seals can partially lift off or even entirely detach during transit dependent on the shipping conditions (heat, air pressure, humidity, etc.). Please place your plate in a corrugated box for maximum protection.

Shipping Address:


MGH CCIB DNA Core Facility
38 Sidney Street / Suite 100
Cambridge, MA 02139
U.S.A.

IMPORTANT: Please note that we are not receiving samples on Saturdays and Sundays or on official holidays.