Software packages are available for all common laboratory computer systems. The packages for personal computers (PC or Macintosh) are able assemble and correct the sequence, those for the larger systems (VAX or Unix) are generally able to analyze the sequence in greater detail. Most laboratories will be able to use sequence assembly programs in their favorite sequence analysis software package. In general, the stages of sequence assembly are gel entry, overlap detection, editing, and reporting. The available programs differ in the ways they handle each of these tasks. No single package is ideal, though all should be adequate for a smaller project such as a single cDNA. Particular attention should be given to the quality and features of the editor, as this is where most time will be spent, and to the possibilities of extending the software to cope with problems that may arise. Good status reports and a choice of methods for overlap detection can save considerable time in resolving ambiguities and correcting errors later. Figure 1 lists some of the commonly used sequence assembly programs. The prices vary widely depending on the features of the package and the options for academic or commercial licenses. Originally, each package used its own “special” codes to represent ambiguous bases and gaps in sequences. Mostpackages now use the standard IUB-IUPAC codes (Figure 2) for the nucleotides, though the program documentation should be checked before starting the project. The task of sequence reading depends on the sequencing protocol used. In many laboratories the sequence is generated on an autoradiograph (Figure 3) from which the sequence is read. Although automated gel readers are on the market, most sequence data is read manually with the aid of a digitizer. Most sequence assembly programs accept DNA sequence read by a sonic digitizer. An example of a device which is supported by most of the available programs is the GrafBar GP-7 [Science Accessories Corporation, Southport, CT, US A and P.M.S. (Instruments) Ltd., Waldeck House, Reform Road, Maidenhead, Berks, SL6 8BX, UK]. Sonic digitizers have a stylus to point to locations on an autoradiograph, which is illuminated from below by a light box.
The Data Evaluation of Interlaboratory Comparisons for Calibration Laboratories
