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In 2006 it was reported that the Largest Ever Killer Crater was found under ice in Antarctica. Gravity measurements taken by NASA's GRACE satellites indicate the crater is 250 million years old.

The size and location of the crater, apparently 480 km across, located in the Wilkes Land region of Eastern Antarctica, south of Australia have lead to suggestion the crater could have initiated the break up of Gondwanaland by creating the tectonic rift that pushed Australia northward. The rift cuts directly through the crater, so the impact may have helped the rift to form. Australia split from the ancient Gondwana super continent about 100 million years ago.

Has the existence of the crater been verified and did it lead to the break up of Gondwanaland and Australia separating from Antarctica?

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To clarify:

Your question is valid, although if you are referring to Dr. von Frese's words as paraphrased in this article, you may be conflating a potential "breakup initiation" with something like the "creation of a geologically weak area prone to rifting under extensional stress". I believe the thought that such a crater could have caused the beginnings of the rifting process was not entertained by Dr. von Frese himself. Instead, according to the article you cited, von Frese explains that Antarctica and Australia began rifting roughly 100 million years ago, and rifting just so happened to occur very near the proposed impact site. From the article:

The rift cuts directly through the crater, so the impact may have helped the rift to form, von Frese said.

It's a little unclear, but I think von Frese is making the connection here that a crater of that size could have potentially created weak planes in the crust near the impact site, which would have directed extensional stress relief to a localized region, thus centralizing a rift locally. This is a plausible conclusion, but as far as I can tell, no evidence other than proximity has been found to support it.

To answer:

The existence of the crater has not been verified, and more recent research casts doubt about the validity of claims that the Wilkes Land structure is in fact an impact crater.

Furthermore, the date for the final breakup of Gondwana is fairly well established, and enough research has been conducted to construct extensive historical summaries of the time period. As stated by von Frese, most geologists put the start of rifting in the Wilkes Land region at about 100 million years ago, in the Cretaceous period.

If we allow the assumption that the crater is indeed caused by a bolide impact, most studies suggest an impact date near the time of the Permian-Triassic boundary, a period known for its multiple mass-extinctions, around 250 million years ago. That would leave roughly 70 million years between impact and Gondwana's very earliest rifting (~180 mya), which is a relatively significant amount of time by tectonic standards. As stated in the above reference, the current most accepted "method" of Gondwanan rifting is via mantle plumes forcing the continents apart from below.

Based solely on these two pieces of evidence, I'd feel relatively confident in putting forth that the breakup of Gondwana was not caused by a bolide impact. If the Wilkes Land structure is ever drilled, and shocked quartz or other impact indicators are found in the samples, I would want to revisit this answer to investigate what I'll call "Dr. von Frese's Planes of Weakness" claim more closely.

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