Their achievement breaks new ground by taking the emerging field of synthetic biology beyond the limits of bacteria and viruses.
Chromosomes are tiny packages of DNA that contain the genes in which the "book of life" is written.
Previously scientists have built artificial versions of bacterial chromosomes but this is the first time the chromosome of a "eukaryote" - a higher organism whose cells contain nuclei, unlike bacteria - has been created.
US synthetic biology pioneer Dr Jef Boeke, director of the New York University Langone Medical Centre, said: "Our research moves the needle in synthetic biology from theory to reality.
"This work represents the biggest step yet in an international effort to construct the full genome of synthetic yeast."
Writing in the journal Science, the team describes how the seven-year project involved piecing together some 273,871 DNA components. The scientists also tweaked nature's design by removing repeated DNA sections thought to be unnecessary to reproduction and growth.