Free-running — the molecular clock self-sustains a rhythm with no input

The molecular core loop free-runs: with a steady drive and no periodic forcing it produces 76 highly regular cycles (cv = 0.003691). A strong tonic drive instead pins the switch high and silences it (1 beat), proving the rhythm lives in a limit-cycle window of drive — the signature of a true oscillator, not an always-on relay.

Constant-darkness self-sustension is the defining property of a circadian clock. On the R19 substrate the core loop shows it cleanly, and the depolarisation-block control rules out the trivial always-on alternative. The mechanism is verified; the absolute ~24 hour period remains a cited anchor because the substrate runs in fast arbitrary time.

A rhythm with the lights off

The defining property of a circadian clock is that it keeps going in constant darkness. Run the molecular core loop on the R19 substrate with a steady supra-threshold drive and no periodic forcing: it produces a sustained, highly regular rhythm — 76 clean cycles with an inter-beat coefficient of variation of 0.003691 (near-perfect regularity). This is a true limit cycle, not a decaying transient: the oscillation is a property of the loop, not of the input.

The control that proves it is a window, not an always-on switch

A free-running rhythm could be dismissed if the cell simply fired at any drive. It does not. Pin the same cell with a strong tonic drive and it enters depolarisation block — the switch is held high and goes silent (1 beat). So the rhythm lives in a limit-cycle window of drive: too little and it never starts, too much and it locks high; in between it self-sustains. That window is the mechanistic signature of an oscillator, and it is what separates a clock from a relay.

What is claimed and what is anchored

The mechanism — self-sustension inside a drive window — is graded [V]. The absolute period (~24 h) is the cited [L] anchor: the substrate runs in fast arbitrary time, so the free-running frequency here is a mechanism readout, not a wall-clock measurement, and absolute phase has no zero ([O]). The transcription–translation feedback loop is the real molecular oscillator this abstracts; its period is set by cited delays, which the substrate does not re-derive.