1. Find sequences of light and heavy chains against the desired antigen (the Ig type doesn't matter:  for heavy chains, IgG, IgA, IgD, ... for light chains, kappa or lambda).
  2. For the heavy chain, replace its constant region with the constant region corresponding to mIgM (membrane-bound IgM).
  3. Both light and heavy chains require leader sequences.  These can be found here: http://www2.mrc-lmb.cam.ac.uk/vbase/alignments2.php  Light and heavy chains can be divided into separate categories based on their variable sequences.  Each of these variable sequences corresponds to a particular leader sequence.  Identify the variable sequences that you are using for the light and heavy chains on the website's list, and then check to see what the corresponding leader sequences are.  If the variable sequence that you have for the light or heavy chain doesn't exactly match any of the variable sequences on the website, choose one that's close and go with that (it could be that it changed due to somatic hypermutation).  Add the leader sequences to the beginning of the variable regions of the light and heavy chains.
  4. Add Q1, Qx, BsaI sites.
  • No labels