The Jacksonville, Tampa and Key West Railway was a railroad and steamboat network in Florida at the end of the 19th century.

The line remains in service today with a vast majority of it now being CSX Transportation's Sanford Subdivision.

The company , by way of congressional land grants thru the general assembly of Florida, by an act of January 6, 1855, 'to provide for and encourage a liberal system of internal improvements in the state,' declared that the lands granted to the state by the acts of congress of March 3, 1845, and September 28, 1850, together with the proceeds thereof, accrued or that might thereafter accrue, should be set apart and made a separate fund, to be called the internal improvement of the state; and that, for the purpose of assuring a proper application of the fund for the objects mentioned, the lands, and the funds arising from the sale thereof, after paying the necessary expenses of selection, management, and sale, should be vested in five trustees, to wit, in the governor of the state, the comptroller of public accounts, the state treasurer, the attorney general, and the register of state lands, and their successors in office, to hold the same for the uses provided in the act; and by its twenty-ninth section, the general assembly reserved the right to grant to such railroad companies, thereafter chartered, as they might deem proper, upon their compliance with the provisions of the act as to the manner of constructing the road and the drainage of the land, the alternate sections of the 'swamp and overflowed lands' for six miles on each side of the line of the road of any such company.

That the legislature of Florida, by act of March 4, 1879, granted to that company alternate sections of the lands given to the state by the act of congress of September 28, 1850, within six miles on each side of the track or line of its road, provided that the company should comply with the specified provisions of the act of January 6, 1855; and further granted to the company, in consideration of the greatly improved value which would accrue to the state from the construction of the road, 10,000 acres of the same class of lands for each mile of road it might construct, such lands to be of those nearest to the line of the road, its branches and extensions,—this last-named grant being made subject to the rights of all creditors of the internal improvement fund, and to the trusts to which the fund was applicable under the act of January 6, 1855.

That on the 27th of June, 1881, the Tampa, Peace Creek & St. John's River Railroad Company, by a resolution of its board of directors, changed its corporate name to Jacksonville, Tampa & Key West Railway Company, and on the 23d of August, 1881, filed a plat of its route with the trustees of the internal improvement fund; and, on the 1st of September, 1881, the trustees passed a resolution reserving from sale for the benefit of the company the even-numbered sections of land for six miles on each side of its line; and, again, on the 21st of September, 1881, acting under the provisions of the act of the legislature of March 12, 1879, 'to amend section 26 of the act 'to provide a general law for the incorporation of railroads and canals,' and to grant aid to railroads and canals incorporated under said act,' they passed a resolution to reserve from sale, to further aid in the construction of the road, a quantity of land in the even-numbered sections within twenty miles of said road sufficient to supply the deficiency existing in the even-numbered sections within six miles of the road.

The Jacksonville, Tampa and Key West Railway then leased the Atlantic Coast, St. Johns and Indian River Railroad, and purchased all of its rolling stock.

The Jacksonville, Tampa and Key West Railway operated the Enterprise to Titusville line as its Indian River Branch.

The Jacksonville, Tampa and Key West Railway remains in service and is today part of CSX's A Line.