Engineering the ribosomal DNA in a megabase synthetic chromosome

Science. 2017 Mar 10;355(6329):eaaf3981. doi: 10.1126/science.aaf3981.


We designed and synthesized a 976,067-base pair linear chromosome, synXII, based on native chromosome XII in Saccharomyces cerevisiae SynXII was assembled using a two-step method, specified by successive megachunk integration and meiotic recombination-mediated assembly, producing a functional chromosome in S. cerevisiae. Minor growth defect "bugs" detected in synXII, caused by deletion of tRNA genes, were rescued by introducing an ectopic copy of a single tRNA gene. The ribosomal gene cluster (rDNA) on synXII was left intact during the assembly process and subsequently replaced by a modified rDNA unit used to regenerate rDNA at three distinct chromosomal locations. The signature sequences within rDNA, which can be used to determine species identity, were swapped to generate a Saccharomyces synXII strain that would be identified as Saccharomyces bayanus by standard DNA barcoding procedures.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Nucleus / genetics
  • Cell Nucleus / ultrastructure
  • Chromosomes, Artificial, Yeast / chemistry*
  • Chromosomes, Artificial, Yeast / genetics
  • Chromosomes, Artificial, Yeast / ultrastructure
  • DNA, Ribosomal / genetics*
  • Genetic Engineering / methods*
  • Genome, Fungal*
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / ultrastructure
  • Synthetic Biology / methods*
  • Transcriptome


  • DNA, Ribosomal