In the most significant blow yet to the Obama administration’s vision of a national high-speed rail network, Gov. Rick Scott of Florida on Wednesday rejected plans for a high-speed link between Tampa and Orlando, in the process turning down more than $2 billion in federal money.

Mr. Scott is the third newly elected Republican governor to turn down a portion of the administration’s national rail system, joining John Kasich of Ohio and Scott Walker of Wisconsin. Each of the three replaced governors who had lobbied for the funds.

Mr. Scott’s move comes a little more than a week after Vice President Joseph R. Biden Jr. called for spending $53 billion on passenger trains and high-speed rail projects over the next six years as part of the administration’s goal of making high-speed rail accessible to 80 percent of Americans within 25 years.

The 85-mile Tampa-to-Orlando segment, on which trains would travel as fast as 170 miles per hour, was to be the showpiece of that initiative — in part because the government already owned much of the right-of-way along the route, which would allow it to be built relatively quickly, and because the fast-moving train would contrast with slow-moving traffic along Interstate 4.