User manual

What is AIMS and what is AIMS in silico software for.

Amplification of intermethylated sites (AIMS) is a powerful tool for differential methylation screening of genomes. Its applications have nevertheless been limited until recently for the absence of systemic approach to AIMS experimental design and of appropriate computer software for the analysis of AIMS results. We have developed AIMS in silico computer suggestion tool capable of predicting possible experimental outcomes, which assists in designing AIMS experiments depending on the research aims and available instrumentation, and in analyzing experimental results from the point of view of genomic locations of the DNA fragments under study.

AIMS in silico computer software is intended to standardize AIMS applications and to turn it into one of the principal approaches towards cancer methylomes studies as well as towards diagnostics in oncology, including early screening.

How to download and install AIMS in silico.

To download AIMS in silico go to http://epigenetic.narod.ru/AimsInSilico/Download.html. Choose “Click here” to download AIMS in silico setup program. Choose a destination to save the setup program on your computer. The setup program is archieved as AimsInSilico.zip file.

Unpack the archieve and run setup.exe. The installer will guide you through the steps required to install AIMS in silico on your computer. You will find the AIMS in silico executable file in the Programs menu, in the RCMG folder.

The program stores data in Microsoft SQL Server database. Microsoft SQL Server Express Edition is required. Available at http://www.microsoft.com/express/sql for free downloading.

Input/output.

This computer program interactive interface invites a user to input the files containing sequences to be analyzed and to select the preliminary experimental conditions, such as restriction endonuclease recognition and cleavage sites, lengths of the adaptor’s shorter and longer components, and/or nucleotide sequences of specific extenders of the universal primer. There also exists an option to limit the lengths of the resulting AIMS fragments, in dependence upon the technique used for their physical separation (as a rule, polyacrylamide gels provide reasonably fine separation between dozens of bases and about a thousand of bases).

The output information includes total number of the PCR products obtainable with AIMS with the preset conditions, distribution of the products by their lengths, and 25-base individual sequences flanking each of them. Graphically information is presented as a virtual capillary electrophoregram drawn as a superposition of Gaussian peaks each corresponding to an individual AIMS product. The peaks are clickable, calling forth their individual information in a separate window.

Flanking 3’- and 5’- nucleotide sequences suggested by AIMS in silico computer program for each PCR product represent unique genomic descriptors that can be used to align DNA fragments under study to the GenBank genome database by BLAT software.

How to handle.

Run AIMS in silico program by following the path: Start – Programs – RCMG - AIMS in silico.

The following window will be presented:

Add file(s) with the genome sequences to analyze by clicking the “Add file” button. Enter restriction site (e.g., C^CCGGG) into the “Restriction site” window. Specify fragment length threshold. If you want to disable “Fragment length threshold” option, unselect the corresponding box.

After the genome sequence has been entered and the analysis parameters have been set, click “Go” to start analysis. This will create a database table containing the descriptions of the digestion products obtainable under selected conditions.

It may take up to an hour, depending on the machine, to perform this initial analysis for the gross genomes, like the human genome. Nevertheless, after the digest has been saved in a database table, every further query is operated in virtually no time.

The progress of the genome analysis may be followed in the “Progress” window.

In order to save time for the researchers using SmaI/XmaI isoschizomers (original AIMS protocol) our software contains a built-in database table of a human genome digest with a restriction enzyme recognizing CCCGGG sequences, with the length threshold of 1kb. It can be found in the “Restriction product” menu as “C^CCGGG Length: 0..1000”.

Choose this table or the one you created of your own design. Input the desired primer extender, e.g., CСG, and the shorter and the longer adapter components lengths.

Click the “Query” button. A simulation of CE chromatogram will appear, supplemented with the following information: primer extender (ccg), total count of the predicted resulting fragments (74), numbers of fragments of the same lengths in nucleotides.

The peaks are clickable. Enlarge the desired area and click the peak of interest. You will be presented the information on the length of the selected AIMS product, the number of products of the same size, and the flanking sequence identifiers of each product.