…which cannot be worked or planted… (Deuteronomy 21:4)
The ceremony of the eglah arufah – the decapitated calf, as described in the previous mitzvah – was held at a river. After the ceremony, the land where the calf was beheaded is not permitted to be worked. Ever.
The reason for this ban is related to the reason for the eglah arufah ceremony in the first place. The ceremony was held to arouse the people against such evils as the murder that occurred in their own back yard. The fact that the spot where the calf was slaughtered could never be planted or plowed served as an eternal reminder of this message.
The Sefer HaChinuch asks an excellent question: people don’t typically do a lot of farming at the river, so what’s the point of forbidding something that isn’t going to be done anyway? The Sefer HaChinuch has an equally excellent answer: there are plenty of dried riverbeds that are planted. Even if the river where an eglah arufah ceremony is held runs dry and becomes suitable for planting, it still may not be done.
Even though the eglah arufah ceremony is only held when the Temple is standing, the ban on working the place where it was held is perpetual. Even today, if there was a place that we knew such a ceremony to have been held, we would not be permitted to work that site.
This mitzvah is discussed in the Talmud in tractate Sotah on page 47b. It is codified in the Mishneh Torah in the tenth chapter of Hilchos Rotzeiach. This mitzvah is #309 of the 365 negative mitzvos in the Rambam’s Sefer HaMitzvos; it is not listed in the Sefer HaMitzvos HaKatzar of the Chofetz Chaim.