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Hello Ms. Brook,
The National Archives and Records Administration holds permanently valuable records of the federal government. Since railroads are private entities, albeit ones with significant mingling with government, the chances of NARA holding any records of your ancestors’ employment experience are quite slim.
Had any of your ancestors been employed by a railroad after 1936 there may be a record of pensions or related materials with the Railroad Retirement Board.
Fortunately railroads are a high interest research topic and there are many other resources at your disposal.
This familysearch Webpage provides some basic information. It recommends consulting railroad directories that may include your ancestors if they were senior enough. Another method would be to contact railroad company records repositories. These collections are not in a single place and their titles may only reflect the railroad companies’ name at the time the record were deposited rather than what it was historically.
Here is another online resource:
http://www.genealogytoday.com/guide/railroad-employees.html : List of Short Lines across America
The Library of Congress and the National Archives both have extensive railroad map collections. Please see this LoC page and please email firstname.lastname@example.org to begin the identification process.
We hope these resources provide a helpful start. Best of luck with your research!
Compiled by Alex Champion