Searched for James Knowlton on FamilySearch, but did not find a matching profile. 23 Mar 2021.
Sources
Mayflower Families Through Five Generations Vol. 15 p. 163
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DNA Connections
It may be possible to confirm family relationships with James by comparing test results with other carriers of his ancestors' Y-chromosome or mitochondrial DNA.
Y-chromosome DNA test-takers in his direct paternal line on WikiTree:
It is likely that these autosomal DNA test-takers will share some percentage of DNA with James:
~0.39%David Smith :
23andMe, yourDNAportal DAVa1bce514
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AncestryDNA, yourDNAportal DAVa1bce514, Ancestry member d__smith
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Family Tree DNA Family Finder, yourDNAportal DAVa1bce514, FTDNA kit #178866