Science - USA (2022-02-25)

13,513,873 sites contain at least two mutations
affecting more than one sample, which implies
that up to 17.5% of variable sites could result
from more than one ancestral mutation. A high
proportion of sites with more than∼100 muta-

tions on chromosome 20 have sequencing or

alignment quality issues as defined by the

TGP accessibility mask ( 6 ) or are in minimal

linkage disequilibrium to their surrounding

sites (fig. S6), which suggests that they are

largely erroneous. Moreover, analysis of data

simulated with an empirically calibrated error

profile and evaluation of the enrichment of

multiple mutations at sites with known ele-

vated mutation rates suggests that most of

Wohnset al.,Science 375 , eabi8264 (2022) 25 February 2022 2of9

A = mutation

Relative age

B

Infer Tree

Sequence Topology

Order by

Frequency

Constrain Ages with

Ancient Samples (if available)

Older

Older

Order by

Estimated Age

Step 0

Step 1

Date Tree

Sequence

Step 2

Step 3

Step 4

Modern Samples Only

Modern +

Ancient Samples (if

available)

C

CEU CHB YRI

D

Ancient Sample

5

6

4

0 1 2 3

7

6

4

0 1 2 3

5

7

6

4

0 1 2 3

Fig. 1. Schematic overview and validation of the inference methodology.
(A) An example tree sequence topology with four samples (nodes 0 to 3),
two marginal trees, four ancestral haplotypes (nodes 4 to 7), and two mutations.
Tspanmeasures the genomic span of each marginal tree topology, with
the dotted line indicating the location of a recombination event. The graph
representation is equivalent to the tree representation. (B) Schematic
representation of the inference methodology. Step 0: Alleles are ordered by
frequency (freq.); the mutation represented by the four-point star is considered
to be older. Step 1: The tree sequence topology is inferred withtsinferusing
modern samples. Step 2: The tree sequence is dated withtsdate. Step 3:
Node date estimates are constrained with the known age of ancient samples.
Step 4: Ancestral haplotypes are reordered by the estimated age of their focal
mutation; the five-pointed star mutation is now inferred to be older. The

algorithm returns to step 1 to reinfer the tree sequence topology with ancient

samples. Arrows refer to modes of operation: steps 0, 1, and 2 only (red);

steps 0, 1, 2, 4, 1, and 2 (green); or steps 0, 1, 2, 3, 4, 1, and 2 (blue) ( 24 ).

(C) Scatter plots and accuracy metrics comparing simulated (xaxis) and inferred

(yaxis) mutation ages frommsprimeneutral coalescent simulations, using

tsdatewith the simulated topology (left) and inferred topology fromtsinfer

(right). RMSLE, root mean squared log error. (D) Accuracy metrics, RMSLE (top),

and Spearman rank correlation coefficient (r) (bottom), with modern samples

only (first panel), after one round of iteration (second panel), and with increasing

numbers of ancient samples (third panel) [colored arrows as in (B)]. Ancient

samples from three eras of human history are considered, as in the schematic

( 24 ). CEU, Utah residents with Northern and Western European Ancestry;

CHB, Han Chinese; YRI, Yorubans.

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