GeneArt online portal tutorial

Transcriptie

1 GeneArt online portal tutorial Part 3: How to order GeneArt Strings DNA Fragments and Libraries

2 Introduction This tutorial is intended to help new users with placing an order for Invitrogen GeneArt services using our GeneArt online portal. Part 3 guides you on how to order GeneArt Strings DNA Fragments and Libraries. Video tutorials are also available at thermofisher.com/genearttutorials

3 Contents 1. GeneArt Strings DNA Fragments and Libraries How to set up an order for GeneArt Strings DNA Fragments How to use the GeneArt Strings DNA Fragments assembly assistant 8 2. Contacts for GeneArt services 14

4 1. GeneArt Strings DNA Fragments and Libraries 1.1 How to set up an order for GeneArt Strings DNA Fragments 1 2 To set up an order for GeneArt Strings DNA Fragments, drag the Strings DNA Fragmts & Libs icon (1) onto the drop area (2). 4

5 3 4 The red exclamation mark (3) indicates that more information is needed. Click the GeneArt Strings DNA Fragments icon to open the form (4). 5

6 a 8b 9 10 The GeneArt Strings DNA Fragments form has three main tabs: the Enter Sequence tab (5), the Edit Sequence tab (6), and the Optimize Sequence tab (7). The Enter Sequence tab allows you to enter the sequence and provide all basic information. Choose your required service (8a) and your sequence type (8b). For Strings DNA Fragments you can choose to enter a DNA or protein sequence. This option is not available for Strings DNA Libraries. Enter your sequence name (9) and paste your sequence into the tool (10). If you enter a protein as an amino acid sequence, you must optimize it for your desired expression host. Please note that GeneArt Strings DNA Fragments up to 3,000 bp can be produced. Longer sequences need to be ordered as gene synthesis projects (recommended) or you must assemble your gene from several GeneArt Strings DNA Fragments that are <3,000 bp. If you entered a sequence >3,000 bp, a message box will be displayed with the option to shorten the sequence or open the assembly assistant for Strings DNA Fragments. This assistant can help you design your genes to assemble from GeneArt Strings DNA Fragments of <3,000 bp using Invitrogen GeneArt Seamless Cloning and Assembly technology. A description of the GeneArt Strings DNA Fragments assembly assistant is given in section 1.2 below. Editing and optimizing your gene works the same way as for full gene synthesis. For details, see GeneArt online portal tutorial part 2 section 1.3, Editing and optimizing a sequence. The Sequence Summary tab (11) displays an overview of the actual sequence and all edits made. The sequence and the summary can be downloaded. For more details, see GeneArt online portal tutorial part 2 section 1.4, Sequence summary. If you have entered all information, save and close the form (12). 6

7 14 13 The green check mark status indicator should now appear (13). If you are ready to place your order, click Proceed to summary (14). The next step is to add the project to your cart and proceed to checkout. Please read the Adding a project to cart and placing your order tutorial in part 1 chapter 2 of the GeneArt online portal tutorial to see how this process works. 7

8 1.2 How to use the GeneArt Strings DNA Fragments assembly assistant 15 Note that GeneArt Strings DNA Fragments up to 3,000 bp can be produced. Once you have entered a sequence longer than 3,000 bp into the sequence field (10), a message box will be displayed with the option to shorten the sequence or open the assembly assistant for GeneArt Strings DNA Fragments. This assistant can help you with the design of GeneArt Strings DNA Fragments to assemble your >3,000 bp gene using GeneArt Seamless Cloning and Assembly technology. Use the button (15) to start the assembly assistant. 8

9 16 Carefully read the notes for using the assembly assistant, and download and read the description (16) for more information on how to use GeneArt Strings DNA Fragments for assembly of larger genes. Editing and optimizing your sequence works the same way as for gene synthesis. For details, see GeneArt online portal tutorial part 2 section 1.3, Editing and optimizing a sequence. 9

10 Use the Edit tab (17) to indicate the open reading frame (generally recommended; mandatory if gene optimization shall be done). Use the Optimize tab (18) to select an organism for optimization and click the Optimize button. If you intend to preclone your GeneArt Strings subfragments prior to assembly into an intermediate vector using restriction enzymes, make sure to avoid the corresponding recognition sites in these subfragment sequences (19). 10

11 On the Split tab (20), read the information, enter the vector (21) you want to insert your final gene into, and enter exactly 20 nucleotides upstream and downstream of the integration site of the vector (22). Note that you must do the seamless assembly yourself, and we need the information only to prepare the vector file to show you how the final construct will look. If you want to preclone the GeneArt Strings subfragments into an intermediate vector before assembly in the final vector, please select your restriction sites used for precloning, otherwise select None (23). Start the calculation of the fragments by clicking on the Start Split Process button (24). 11

12 On the Sequence Summary tab, carefully read the Important customer information section (25) and scroll down to verify the full gene sequence, including insertion site, gene optimization information, plasmid map, sequence of all GeneArt Strings DNA Fragments, and amplification primers. You can download a.gb file of the final plasmid, the sequences in FASTA format, and a project report for your reference (26). Once all information is correct, click the Create Fragments button (27) to create the fragments and proceed to order. Scroll down in the Sequence Summary tab to see additional information about your project. 12

13 29 28 In the Project Configurator you will now find the GeneArt Strings DNA Fragments needed to assemble your gene of >3,000 bp (28) using the GeneArt Seamless Cloning and Assembly technology. To order, click the Proceed to summary button (29). Please read the Adding a project to cart and placing your order tutorial in part 1 chapter 2 of the GeneArt online portal tutorial to see how this process works. 13

14 2. Contacts for GeneArt services For further information or if you have questions, contact us at Europe: Phone: +49 (0) Fax: +49 (0) America and Asia: Phone: , option 4/4/1 Headquarters 81 Wyman St. Waltham, MA Phone Toll Free in the USA thermofisher.com International Sales For our office locations, please call the division headquarters or refer to our website at thermofisher.com 14

15 15

16 DISCLAIMER: LIFE TECHNOLOGIES AND/OR ITS AFFILIATE(S) DISCLAIM ALL WARRANTIES WITH RESPECT TO THIS DOCUMENT, EXPRESSED OR IMPLIED, INCLUDING BUT NOT LIMITED TO THOSE OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE, OR NON-INFRINGEMENT. IN NO EVENT SHALL LIFE TECHNOLOGIES AND/OR ITS AFFILIATE(S) BE LIABLE, WHETHER IN CONTRACT, TORT, WARRANTY, OR UNDER ANY STATUTE OR ON ANY OTHER BASIS FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT, INCLUDING BUT NOT LIMITED TO THE USE THEREOF. Find out more at thermofisher.com/geneart For Research Use Only. Not for use in diagnostic procedures Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. CO