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Area of Science:

  • Genetics and heredity
  • Molecular biology
  • Gene editing technologies

Background:

  • CRISPR-based gene drives utilize Cas9 and guide RNA (gRNA).
  • Gene drives can be configured as self-contained full gene drives (fGD) or split gene drives (sGD) for enhanced control.
  • Previously engineered split systems can be converted into autonomous full drives.

Purpose of the Study:

  • To examine dual split gene drive systems engineered for conversion into autonomous full drives.
  • To investigate the insertion of these dual systems at the spo11 locus, recoded to restore gene function and organismic fertility.
  • To analyze the kinetic behaviors and transmission efficiencies of split versus full gene drive elements.

Main Methods:

  • Engineering and insertion of dual split gene drive systems at the spo11 locus.
  • Recoding transgenes to restore gene function and organismic fertility.
  • Conducting single generation crosses and multigenerational cage studies to assess drive kinetics and transmission efficiency.

Main Results:

  • Split gene drive (sGD) and full gene drive (fGD) elements showed minimal differences in transmission efficiency in single crosses.
  • The reconstituted spo11 fGD cassette exhibited slower initial drive kinetics compared to the unlinked sGD element in multigenerational studies.
  • Both sGD and fGD elements eventually achieved similar levels of final introduction, despite kinetic differences.

Conclusions:

  • Unexpected kinetic differences between sGD and fGD elements were observed.
  • These kinetic behaviors are likely attributed to transient fitness costs in individuals co-inheriting Cas9 and gRNA transgenes.
  • Split gene drive systems offer potential for greater control, with full gene drives showing distinct kinetic profiles during spread.