Order Form by Levone

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									MBSU DNA Sequencing Service
Please Reply to: Joseph Black Building, University of Glasgow, University Avenue, Glasgow, G12 8QQ, Scotland Telephone: 0141-330-6212. Fax: 0141-330-4600. E-mail: mbsu@udcf.gla.ac.uk Web: http://www.gla.ac.uk/ibls/ASU/MBSU.index.html

Order Form – Reaction to be done by MBSU
Name: Address and Group Leader: Type of DNA: plasmid q PCR product q If PCR product: already cleaned q
S/N Sample Name Vol sample

Date: Tel and e-mail: Keep Samples : Yes q No q

to be cleaned by MBSU q
Primer name (3.2 M) Size V/I* (Kb) S/N

If plasmid, plasmid name: Templiphi: Yes q No q
Sample Name Vol sample Primer name (3.2 M) Size V/I* (Kb)

1 2 3 4 5 6 7 8 9 10 11 12
*V/I:

13 14 15 16 17 18 19 20 21 22 23 24 Vector/Insert PLEASE FILL IN ALL INFORMATION Use dGTP BigDyes kit q
(Only for difficult and repetitive templates) Marker used: Indicated a band in the marker and its amount in ng: Volume of DNA loaded on the agarose gel (in l): Further information:

INVOICE DETAILS
Budget Centre and Holder: Fac. Ref: Authorising signature:

FOR MBSU STAFF
No. of samples using BD: No. samples E-S cleaned: No. of samples using dGTP Kit: No. of samples templiphied: Sequencing date: Gel date:

MBSU DNA Sequencing Service
Please Reply to: Joseph Black Building, University of Glasgow, University Avenue, Glasgow, G12 8QQ, Scotland Telephone: 0141-330-6212. Fax: 0141-330-4600. E-mail: mbsu@udcf.gla.ac.uk Web: http://www.gla.ac.uk/ibls/ASU/MBSU.index.html

Rough Guidelines for the use of the MBSU Sequencing Service
1. We have one abi 377 sequencer with 64 lanes available. 2. We do accept sequencing reactions prepared by customers. In that case the chemistry used MUST BE indicate on the Order Form – Ready reactions (ready-seq.rtf). 3. We expect the customers to provide us with DNA prepared by one of the following methods:  QIASpin mini preps (QIAGEN) of 3-5 ml cultures eluted in 30 l EB buffer  other kind of mini preps: eg. Wizard (please use a lower volume for the elution step: 50 l)  preparation using CsCl (please contact Giorgia before starting the preparation)  Before staring the preparation of Midi and Maxi please contact Giorgia. 4. PCR products have to be cleaned up from any residues from the PCR reactions by cutting the band out from an agarose gel and then extract it using a kit, by PEG precipitation or by PCR cleaning columns. A test gel has to be run after the cleaning procedure to determine the correct amount of DNA recovered. The amount of DNA required for a sequencing reaction varies from 80 ng up to 400-600 ng depending on the size of the PCR product (for more details get in touch with our staff). 5. For quantification purposes ONLY, plasmid DNA must be linearised, or digested, and run on an agarose gel together with a suitable marker. No responsability will be taken by MBSU staff for samples provided using a spectrophotometer as method of estimation. The volume of plasmid DNA and the amount of marker loaded on the gel have to be supplied together with the picture. Please indicate a band of the marker on the gel and write the correspondent amount in ng on the form. Preferred markers are 1Kb plus and 1Kb ladders (Life Technologies) available from IBLS lower campus store. The minimum amount of plasmid dsDNA to provide per reaction is 600-800 ng (half of this amount is required for ssDNA). Unless you have ticked “YES” in the “Keep DNA” box all samples will be discarded as soon as the sequence data has been loaded on the server. Samples kept will be stored in the –20 freezer in boxes labelled with the initials of the customers for a maximum of one month. When the box is full, the MBSU staff will proceed to discard the older samples. 6. We provide common primers such as M13 forward and reverse, T3, T7 and Sp6; if you would like us to use a custom primer it must be provided at a concentration of 3.2 M. Primers will be used at 3-4 pmol/reaction. 7. Samples provided without picture, or low quality picture, will not be processed. We routinely run the 377 for sequencing samples (better sequence quality and longer reads) using Big Dyes ® chemistry. For templates containing repetitive sequences (i.e. GT, CAG or microsatellites) and long G stretches we can provide a dGTP BD kit that could solve sequencing difficulties (In that case please tick the appropriate box on the form). We also have available Dye ET Terminator ®, Dye Terminators® and Dye Primers® chemistries on request. Using the 377 we normally expect to provide our customers with 400-500 bp of good sequence, and could provide up to 700 bp, depending on the quality of DNA supplied. If you need any further information or guidance, feel free to contact us. We aim to have results available in approximately 3-5 working days. Dr. rer. nat. Giorgia R. Riboldi-Tunnicliffe Manager


								
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