Copyright © 2008, CMB
Natalie Kolber, CMB intern for 2014-2015
Click the 'Protocols' button for info on how to set up a sequencing reaction and some product recommendations.
Once you have performed and purified your sequencing reactions, we will run your purified sequencing samples using the Applied Biosystems 3130xl Genetic Analyzer. You need to follow the rules listed below to submit your samples and retrieve your data.
The current cost for each ready-to-sequence sample is $1.50. It is relatively low because the reagents are kindly supplied by Prof. Steven Williams’ lab for now. It may change in the future.
- Each lab is responsible for purification of your DNA sequencing samples before submitting to the CMBS.
- Put PI initials plus sample’s serial number (must be 4 digits unless you’ve passed 9999, then use 5 digits) on the top of each tube containing to-be-sequenced sample, such as “LAK0123”.
- Every time you submit a new batch of samples, the serial number of the sample must be sequential from the last batch.
- There are two boxes in the freezer (on the side wall) in Ford 123. They are labeled as “Sequences To Be Run on 3100 Box 1” and “Box 2”. Put 8 samples in a row only. Put samples in numerical order. Please fill out the Box 1 before you put into Box 2.
- Fill out the order form which is in the binder on the counter across from the freezer.
- The CMBS will run sequencing samples on the “first come, first served” basis.
- We are using the ABI3130xl DNA Genetic Analyzer to run our sequencing samples. Because 16 samples are injected to the capillaries simultaneously, it is better to try to submit several sequences at a time so we can try to fill all slots. Samples will generally be loaded onto the machine between 11am and noon every weekday.
- After samples have been run and sequence data have been extracted, the CMBS will transfer the data to your DROPBOX in the SCIENCE.DATAVOL server, usually by 9am the next business day if sequences are in before 11am.
- Each lab should be responsible for having enough room for sequencing drop-off in your DROPBOX.
- Currently the CMBS does not have a second place to store your sequence data. The sequence data will be trashed after a certain time (for now, it’s 3 months) in order to keep the sequencer functioning properly. You should be responsible for your data storage.
We will run your post-PCR samples using the Applied Biosystems 3130xl Genetic Analyzer. The current cost for each sample is $2.00. It may change in the future. Please contact Lou Ann Bierwert directly at email@example.com if you plan to have fragment analysis samples. When ready, the samples should be entered in the sequence order book just like sequences in the appropriate section pertaining to your class or lab. Simply write "HID" with your sample serial numbers. Lou Ann will run the samples for you and show you how to analyze the data and print a plot for each sample. This print out will be your only product. The new software no longer allows you to align the various profiles for comparison. In general, a power point of the pdfs of the appropriate profiles is what is used for presentation to a class.
We also provide the instruments and training for the following experiments. You need to contact Lou Ann Bierwert at firstname.lastname@example.org to get training and make areservation before usage. Click here for more information on these instruments.
PCR: MJ PTC-200 Thermal Cycler
Life Technologies Veritii Thermal Cycler
Real-Time PCR: Applied Biosystems 7300 Real-Time PCR System
Life Technologies StepOne Plus Real-Time PCR System
Microarray Scan: GenePix Personal 4100A Scanner
Spectramax M5 Plate Reader
Beckman DU 530 Spec
Spectramax M5 Plate Reader
Centrifuge: Tritech Refrigerated Centrifuge - 15ml, 50ml, and plates
Eppendorf centrifuge 5810
Freezer: 2 X Thermo Fisher Corporation Revco -20C Freezer
Gel Imaging: Amhersham Biosciences Typhoon 9200 Variable Mode Imager
Bio-Chemi darkroom cabinet and imager
PrepOne Sapphire Blue Light Illuminator and Image Capture System