Spotlight

A showcase of research and scholarship

in selected articles from 2017
2017/18
Editorial Board ASSOCIATE EDITORS
Robert R. H. Anholt
EDITOR IN CHIEF North Carolina State
University
Brenda Andrews
University of Toronto Michael J. Axtell
Pennsylvania State
University
DEPUTY EDITOR
Danika L. Bannasch
Dirk-Jan de Koning University of California,
Swedish University of Davis
Agricultural Sciences
Arash Bashirullah
University of
EXECUTIVE EDITOR Wisconsin-Madison
Tracey DePellegrin Judith Berman
University of Minnesota
MANAGING EDITOR & Tel Aviv University
Ruth Isaacson James A. Birchler
University of Missouri
ASSISTANT EDITOR Charles Boone
Sarah Bay University of Toronto
Michael Boutros
SENIOR EDITORS DKFZ & University
of Heidelberg
Eduard Akhunov
Kansas State University Patrick J. Brown
University of
Katrien M. Devos California, Davis
University of Georgia
Rita M. Cantor
Susan L. Forsburg University of
University of California, Los Angeles
Southern California
Susan E. Celniker
Rob J. Kulathinal Lawrence Berkeley
Temple University National Laboratory
Howard D. Lipshitz Aravinda Chakravarti
University of Toronto Johns Hopkins
Jeffrey Ross-Ibarra University School
University of of Medicine
California, Davis J. Michael Cherry
Stanford University
SERIES EDITOR Timothy J. Close
Lauren M. McIntyre University of
University of Florida California, Riverside
Barak A. Cohen
Washington University
School of Medicine
Josep M. Comeron
University of Iowa
Gloria M. Coruzzi
New York University
William S. Davidson
Simon Fraser University
Kelly Dawe
University of Georgia
Gustavo A. Ross Houston Chad L. Myers Lars M. Steinmetz
de los Campos The Roslin Institute University of Minnesota European Molecular
Michigan State Emma Huang Biology Laboratory &
Brian Oliver
University Janssen Pharma R & D Stanford University
NIDDK, National
Job Dekker Timothy R. Hughes Institutes of Health Hidenori Tachida
University of University of Toronto Kyushu University
Fernando
Massachusetts Scott A. Jackson Pardo-Manuel Kevin Thornton
Medical School University of Georgia University of
de Villena
Rebecca W. Doerge University of North California, Irvine
Mattias Jakobsson
Purdue University Uppsala University Carolina at Chapel Hill David W. Threadgill
School of Medicine Texas A&M University
Andrew Doust Jean-Luc Jannink
Oklahoma State USDA-ARS Isobel Parkin Sarah A. Tishkoff
University Agriculture and University of
Sue L. Jaspersen Agri-Food Canada
Aimée M. Dudley Pennsylvania
Stowers Institute for
Pacific Northwest Medical Research Andrew H. Paterson Olga Troyanskaya
Diabetes Research University of Georgia Princeton University
Institute Nicholas Katsanis
Duke University Craig S. Pikaard Mike Tyers
Jay C. Dunlap Indiana University Université de Montréal
Dartmouth John K. Kim
Johns Hopkins James Prendergast Joshua Udall
Medical School
University University of Edinburgh Brigham Young
Mark Estelle University
Yuseob Kim Bruce Reed
University of
Ewha Womans University of Waterloo Marian Walhout
California, San Diego
University Jasper Rine University of
Justin D. Faris Massachusetts
Éric Lécuyer University of
USDA-ARS Cereal Medical School
Institut de recherches California, Berkeley
Crops Research Unit
cliniques de Montréal Antonis Rokas Mick Watson
David S. Fay (IRCM) University of Edinburgh
Vanderbilt University
University of Wyoming
Siu Sylvia Lee Matthew S. Sachs Jonathan F. Wendel
Justin C. Fay Cornell University Iowa State University
Texas A&M University
Washington
University in St. Louis Jianxin Ma Helen K. Salz Randall Wisser
Purdue University Case Western Reserve University of Delaware
Elizabeth R. Gavis
Stuart J. Macdonald University Dani Zamir
Princeton University
The University Michael J. Scanlon The Hebrew University
Cayetano Gonzalez of Kansas of Jerusalem
Cornell University
IRB Barcelona
Trudy Mackay David S. Schneider Monique Zetka
Brian D. Gregory North Carolina McGill University
Stanford University
University of State University
Pennsylvania Robert A. Sclafani
Christian R. Marshall University of Colorado
David J. Gresham The Hospital for School of Medicine
New York University Sick Children
Steve Scofield
Erich Grotewold Sarah Mathews USDA-ARS
The Ohio State The Commonwealth
University Tanja Slotte
Scientific and
University of
David J. Grunwald Industrial Research
Stockholm
The University of Utah Organisation (CSIRO)
Shavannor M. Smith
Kris Gunsalus Andrew S. McCallion
The University
New York University Johns Hopkins
of Georgia
University School
Jay R. Hesselberth
of Medicine Marcus B. Smolka
University of Colorado
Cornell University
School of Medicine Kim S. McKim
Rutgers University Dina A. St. Clair
Charles S. Hoffman
University of
Boston College Todd Mockler
California, Davis
Donald Danforth Plant
James B. Holland
Science Center
USDA & North Carolina
State University
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City University of New York

ON THE COVER A male orchid bee (Euglossa dilemma) drinking nectar. Whole genome
sequencing efforts by Brand et al. revealed that E. dilemma has one of the largest genomes
2 known for insects. Photo: Thomas Eltz.
Why submit your work to G3? We publish papers
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Brenda Andrews
Editor-in-Chief,
G3: Genes | Genomes | Genetics
3
 GENOME REPORT

The Nuclear and Mitochondrial Genomes

of the Facultatively Eusocial Orchid Bee
Euglossa dilemma
Philipp Brand, Nicholas Saleh, Hailin Pan, Cai Li, Karen M. Kapheim,
and Santiago R. Ramírez
G3: Genes | Genomes | Genetics September 2017, 7: 2891–2898

EDITORS’ NOTE Bees provide indispensable pollination services and have

become important models for the study of learning and memory, plant–insect
interactions, and social behavior. Here, Brand et al. describe the nuclear
and mitochondrial genomes of the facultatively eusocial orchid bee Euglossa
dilemma. This study provides valuable genomic resources for genetic
analyses on the ecology, evolution, and conservation of orchid bees and
the evolution of sociality in bees.

ABSTRACT Bees provide indispensable pollination services to both

agricultural crops and wild plant populations, and several species of bees
have become important models for the study of learning and memory,
plant–insect interactions, and social behavior. Orchid bees (Apidae:
Euglossini) are especially important to the fields of pollination ecology,
evolution, and species conservation. Here we report the nuclear and
mitochondrial genome sequences of the orchid bee Euglossa dilemma
Bembé & Eltz. E. dilemma was selected because it is widely distributed,
highly abundant, and it was recently naturalized in the southeastern United
States. We provide a high-quality assembly of the 3.3 Gb genome, and an
official gene set of 15,904 gene annotations. We find high conservation of
gene synteny with the honey bee throughout 80 MY of divergence time.
This genomic resource represents the first draft genome of the orchid bee
genus Euglossa, and the first draft orchid bee mitochondrial genome, thus
representing a valuable resource to the research community.

4
 MUTANT SCREEN REPORT

Genome-Wide Screen for Haploinsufficient

Cell Size Genes in the Opportunistic Yeast
Candida albicans
Julien Chaillot, Michael A. Cook, Jacques Corbeil, and Adnane Sellam
G3: Genes | Genomes | Genetics February 2017, 7: 355–360

EDITORS’ NOTE The Start point is a major cell cycle checkpoint in yeast
that controls commitment to cell division in late G1 phase, and a number
of critical Start regulators are haploinsufficient for cell size. Challiot et al.
used elutriation-barcode sequencing methodology to conduct a quantitative
analysis of size in the opportunistic yeast Candida albicans. They identify
conserved regulators and biological processes that are required to maintain
size homeostasis as well as novel C. albicans-specific size genes, thus
providing a conceptual framework for future mechanistic studies.

ABSTRACT One of the most critical but still poorly understood aspects of
eukaryotic cell proliferation is the basis for commitment to cell division in late
G1 phase, called Start in yeast and the Restriction Point in metazoans. In all
species, a critical cell size threshold coordinates cell growth with cell division
and thereby establishes a homeostatic cell size. While a comprehensive
survey of cell size genetic determinism has been performed in the saprophytic
yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, very
little is known in pathogenic fungi. As a number of critical Start regulators
are haploinsufficient for cell size, we applied a quantitative analysis of the
size phenome, using elutriation-barcode sequencing methodology, to 5639
barcoded heterozygous deletion strains of the opportunistic yeast Candida
albicans. Our screen identified conserved known regulators and biological
processes required to maintain size homeostasis in the opportunistic yeast
C. albicans. We also identified novel C. albicans-specific size genes and
provided a conceptual framework for future mechanistic studies. Interestingly,
some of the size genes identified were required for fungal pathogenicity
suggesting that cell size homeostasis may be elemental to C. albicans fitness
or virulence inside the host.

5
 SOF T WARE AND DATA RESOURCES

rSalvador:
An R Package for the Fluctuation Experiment
Qi Zheng
G3: Genes | Genomes | Genetics December 2017, 7: 3849–3856

EDITORS’ NOTE For nearly 75 years, the Luria-Delbrück fluctuation

experiment has been the preferred method for measuring microbial
mutation rates in the laboratory; however, popular web analysis tools lack
the appropriate computational methodology to properly analyze these assays.
In this article, Qi Zheng describes rSalvador—an R package that addresses
three types of known problems in fluctuation assay analysis.

ABSTRACT The past few years have seen a surge of novel applications of the
Luria-Delbrück fluctuation assay protocol in bacterial research. Appropriate
analysis of fluctuation assay data often requires computational methods that
are unavailable in the popular web tool FALCOR. This paper introduces an
R package named rSalvador to bring improvements to the field. The paper
focuses on rSalvador’s capabilities to alleviate three kinds of problems
found in recent investigations: (i) resorting to partial plating without properly
accounting for the effects of partial plating; (ii) conducting attendant fitness
assays without incorporating mutants’ relative fitness in subsequent data
analysis; and (iii) comparing mutation rates using methods that are in general
inapplicable to fluctuation assay data. In addition, the paper touches on
rSalvador’s capabilities to estimate sample size and the difficulties related to
parameter nonidentifiability.

6
Mutant screen results gathering dust in your lab notebook?
WGS datasets languishing on your hard drive?
New software tools going unshared?

We’ve got an article

format for that.
Mutant Describe results of
Screen mutant screens
Reports

Describe whole
Genome genome sequence
(WGS) data of
Reports organisms and/or
strains

Software Describe novel

software for genetic
& Data data analyses and
Resources database resources

g3journal.org/content/article-types
 INVESTIG ATION

De Novo Genome and Transcriptome Assembly

of the Canadian Beaver (Castor canadensis)
Si Lok, Tara A. Paton, Zhuozhi Wang, Gaganjot Kaur, Susan Walker, Ryan K. C. Yuen,
Wilson W. L. Sung, Joseph Whitney, Janet A. Buchanan, Brett Trost, Naina Singh,
Beverly Apresto, Nan Chen, Matthew Coole, Travis J. Dawson, Karen Ho, Zhizhou Hu,
Sanjeev Pullenayegum, Kozue Samler, Arun Shipstone, Fiona Tsoi, Ting Wang,
Sergio L. Pereira, Pirooz Rostami, Carol Ann Ryan, Amy Hin Yan Tong, Karen Ng,
Yogi Sundaravadanam, Jared T. Simpson, Burton K. Lim, Mark D. Engstrom,
Christopher J. Dutton, Kevin C. R. Kerr, Maria Franke, William Rapley,
Richard F. Wintle, and Stephen W. Scherer
G3: Genes | Genomes | Genetics February 2017, 7: 755–773

EDITORS’ NOTE Lok et al. break ground by reporting the genome of the
Canadian beaver—the first large rodent species and first mammalian genome
assembled directly from uncorrected and modest coverage (30x) noisy long
reads from single molecule sequencing. Further optimization of this approach
is expected to benefit large genomics projects for variant detection at cohort
or population scales. The genome was annotated with over 9,800 full-length
ORFs constructed from the beaver leukocyte and muscle transcriptomes and
provides an important genomic resource for rodent comparative biology.

ABSTRACT The Canadian beaver (Castor canadensis) is the largest

indigenous rodent in North America. We report a draft annotated assembly
of the beaver genome, the first for a large rodent and the first mammalian
genome assembled directly from uncorrected and moderate coverage
(< 30 ×) long reads generated by single-molecule sequencing. The genome
size is 2.7 Gb estimated by k-mer analysis. We assembled the beaver
genome using the new Canu assembler optimized for noisy reads. The
resulting assembly was refined using Pilon supported by short reads (80 ×)
and checked for accuracy by congruency against an independent short read
assembly. We scaffolded the assembly using the exon–gene models derived
from 9805 full-length open reading frames (FL-ORFs) constructed from the
beaver leukocyte and muscle transcriptomes. The final assembly comprised
22,515 contigs with an N50 of 278,680 bp and an N50-scaffold of 317,558 bp.
Maximum contig and scaffold lengths were 3.3 and 4.2 Mb, respectively, with
a combined scaffold length representing 92% of the estimated genome size.
The completeness and accuracy of the scaffold assembly was demonstrated
by the precise exon placement for 91.1% of the 9805 assembled FL-ORFs and
83.1% of the BUSCO (Benchmarking Universal Single-Copy Orthologs) gene
set used to assess the quality of genome assemblies. Well-represented were
genes involved in dentition and enamel deposition, defining characteristics of
rodents with which the beaver is well-endowed. The study provides insights
for genome assembly and an important genomics resource for Castoridae
and rodent evolutionary biology.

8
De novo Genome Assembly of the Canadian beaver (Castor canadensis). The
beaver is an iconic national symbol for the sovereignty of Canada. The “Three
Pence Beaver” was Canada’s first postage stamp, issued in 1851 in recognition
of the beaver as the economic engine that drove the colonial expansion that led
to the founding of Canada. The beaver dam was also symbolic of the young
country building its towns, cities, and communities. The stamp was designed by
Sir Sandford Fleming, a proponent of the worldwide standard time zone, and
Canada’s foremost railway engineer and distinguished scientist of the 19th century.
Photo: CS4 courtesy of MM.

9
 GENOMIC SELECTION

Persistency of Prediction Accuracy and Genetic

Gain in Synthetic Populations Under Recurrent
Genomic Selection
Dominik Müller, Pascal Schopp, and Albrecht E. Melchinger
G3: Genes | Genomes | Genetics March 2017, 7: 801–811

EDITORS’ NOTE Plant breeding techniques such as recurrent selection

are used to improve breeding populations. Here, Müller, Schopp, and
Melchinger simulate synthetic populations by intermating parents from
ancestral populations with different linkage disequilibrium and subjecting
them to 30 cycles of recurrent genomic selection to analyze prediction
accuracy and genetic gain. They examine how variables change when the
number of parents increases and report a high persistency of accuracy.

ABSTRACT Recurrent selection (RS) has been used in plant breeding

to successively improve synthetic and other multiparental populations.
Synthetics are generated from a limited number of parents (Np)1 but little is
known about how Np affects genomic selection (GS) in RS, especially the
persistency of prediction accuracy (rg,ĝ) and genetic gain. Synthetics were
simulated by intermating Np = 2–32 parent lines from an ancestral population
with short- or long-range linkage disequilibrium (LDA) and subjected to
multiple cycles of GS. We determined rg,ĝ and genetic gain across 30
cycles for different training set (TS) sizes, marker densities, and generations
of recombination before model training. Contributions to rg,ĝ and genetic
gain from pedigree relationships, as well as from cosegregation and LDA
between QTL and markers, were analyzed via four scenarios differing in (i) the
relatedness between TS and selection candidates and (ii) whether selection
was based on markers or pedigree records. Persistency of rg,ĝ was high for
small Np1 where predominantly cosegregation contributed to rg,ĝ, but also
for large Np1 where LDA replaced cosegregation as the dominant information
source. Together with increasing genetic variance, this compensation resulted
in relatively constant long- and short-term genetic gain for increasing Np > 4,
given long-range LDA in the ancestral population. Although our scenarios
suggest that information from pedigree relationships contributed to rg,ĝ for
only very few generations in GS, we expect a longer contribution than in
pedigree BLUP, because capturing Mendelian sampling by markers reduces
selective pressure on pedigree relationships. Larger TS size (NTS ) and higher
marker density improved persistency of rg,ĝ and hence genetic gain, but
additional recombinations could not increase genetic gain.

10
 INVESTIG ATION

The Accuracy and Bias of Single-Step Genomic

Prediction for Populations Under Selection
Wan-Ling Hsu, Dorian J. Garrick, and Rohan L. Fernando
G3: Genes | Genomes | Genetics August 2017, 7: 2685–2694

EDITORS’ NOTE When genotypes are observed for all individuals, the
centering of genotype covariates has no effect on genomic prediction; this
is not the case when some of the genotypes are missing. Single-step
genomic prediction is used to combine information from genotyped and
non-genotyped individuals. In this situation, the observed genotypes should
be centered using genotype means from unselected founders, which may
not be available. Here, Hsu, Garrick, and Fernando use computer simulation
to study an alternative analysis that does not require centering genotypes but
instead fits the mean of unselected individuals as a fixed effect.

ABSTRACT In single-step analyses, missing genotypes are explicitly or

implicitly imputed, and this requires centering the observed genotypes
using the means of the unselected founders. If genotypes are only available
for selected individuals, centering on the unselected founder mean is not
straightforward. Here, computer simulation is used to study an alternative
analysis that does not require centering genotypes but fits the mean μg of
unselected individuals as a fixed effect. Starting with observed diplotypes
from 721 cattle, a five-generation population was simulated with sire selection
to produce 40,000 individuals with phenotypes, of which the 1000 sires had
genotypes. The next generation of 8000 genotyped individuals was used
for validation. Evaluations were undertaken with (J) or without (N) μg when
marker covariates were not centered; and with (JC) or without (C) μg when all
observed and imputed marker covariates were centered. Centering did not
influence accuracy of genomic prediction, but fitting μg did. Accuracies were
improved when the panel comprised only quantitative trait loci (QTL); models
JC and J had accuracies of 99.4%, whereas models C and N had accuracies
of 90.2%. When only markers were in the panel, the 4 models had accuracies
of 80.4%. In panels that included QTL, fitting μg in the model improved
accuracy, but had little impact when the panel contained only markers.
In populations undergoing selection, fitting in the model is recommended
to avoid bias and reduction in prediction accuracy due to selection.

11
12
 HERITAGE & INHERITANCE More than two-thirds of the ~10.7 million
Africans who were forcibly brought to the New World ended up in Latin
America. Despite the impact that African descendants have had on the region’s
demography, there have been relatively few studies on the genetic ancestry of
Afro-Latinos. ChocoGen is a collaborative research project aimed at the discovery
and characterization of the genetic heritage of the people of Chocó, a state located
on the Pacific coast of Colombia that has a predominantly African genetic heritage
with admixture from Europe and the Americas. The San Pacho festival shown
here is a celebration of Afro-Colombian heritage and is held every year in the
capital city of Quibdó. Photo: Photography and film archive of the Universidad
Tecnológica del Chocó.

A Comparative Analysis of Genetic Ancestry and Admixture

in the Colombian Populations of Chocó and Medellín
Andrew B. Conley, Lavanya Rishishwar, Emily T. Norris,
Augusto Valderrama-Aguirre, Leonardo Mariño-Ramírez,
Miguel A. Medina-Rivas, and I. King Jordan
G3: Genes | Genomes | Genetics October 2017, 7: 3435–3447 13
 INVESTIG ATION

The Effect of Temperature

on Drosophila Hybrid Fitness
Charles J. J. Miller and Daniel R. Matute
G3: Genes | Genomes | Genetics February 2017, 7: 377–385

EDITORS’ NOTE Drosophila hybrids have been crucial for our understanding
of the genetic basis of hybrid incompatibility. Miller and Matute investigate
the environmental dependence of such incompatibilities by crossing
Drosophila species and measuring the effects of temperature on the fitness
of the hybrid offspring. Their results suggest that hybrid inviability is not solely
the product of genetic interactions in the hybrid offspring; hybrid inviability
must be considered within the broader environmental and organismal context
in which it is observed.

ABSTRACT Mechanisms of reproductive isolation inhibit gene flow between

species and can be broadly sorted into two categories: prezygotic and
postzygotic. While comparative studies suggest that prezygotic barriers
tend to evolve first, postzygotic barriers are crucial for maintaining species
boundaries and impeding gene flow that might otherwise cause incipient
species to merge. Most, but not all, postzygotic barriers result from genetic
incompatibilities between two or more loci from different species, and occur
due to divergent evolution in allopatry. Hybrid defects result from improper
allelic interactions between these loci. While some postzygotic barriers are
environmentally-independent, the magnitude of others has been shown
to vary in penetrance depending on environmental factors. We crossed
Drosophila melanogaster mutants to two other species, D. simulans and
D. santomea, and collected fitness data of the hybrids at two different
temperatures. Our goal was to examine the effect of temperature on
recessive incompatibility alleles in their genomes. We found that temperature
has a stronger effect on the penetrance of recessive incompatibility alleles
in the D. simulans genome than on those in the D. santomea genome.
These results suggest that the penetrance of hybrid incompatibilities can
be strongly affected by environmental context, and that the magnitude of
such gene-by-environment interactions can be contingent on the genotype
of the hybrid.

14
 INVESTIG ATION

Evaluation and Design of Genome-Wide

CRISPR/SpCas9 Knockout Screens
Traver Hart, Amy Hin Yan Tong, Katie Chan, Jolanda Van Leeuwen,
Ashwin Seetharaman, Michael Aregger, Megha Chandrashekhar, Nicole Hustedt,
Sahil Seth, Avery Noonan, Andrea Habsid, Olga Sizova, Lyudmila Nedyalkova,
Ryan Climie, Leanne Tworzyanski, Keith Lawson, Maria Augusta Sartori,
Sabriyeh Alibeh, David Tieu, Sanna Masud, Patricia Mero, Alexander Weiss,
Kevin R. Brown, Matej Usaj, Maximilian Billmann, Mahfuzur Rahman, Michael
Costanzo, Chad L. Myers, Brenda J. Andrews, Charles Boone, Daniel Durocher,
and Jason Moffat
G3: Genes | Genomes | Genetics August 2017, 7: 2719–2727

EDITORS’ NOTE CRISPR-based gene editing approaches have rapidly

evolved to provide an efficient means of manipulating human genome
sequences in laboratory conditions. In fact, gene knockout screens in
human cells are transforming human functional genomics and facilitating drug
target discovery efforts for a long list of disease models. This study from Hart
et al. provides an analysis of the current CRISPR-based genetic screens in
human cell lines and leverages these results to generate an improved, pooled,
genome-scale CRISPR-Cas9 library of more efficient gene knockout screens.

ABSTRACT The adaptation of CRISPR/SpCas9 technology to mammalian

cell lines is transforming the study of human functional genomics. Pooled
libraries of CRISPR guide RNAs (gRNAs) targeting human protein-coding
genes and encoded in viral vectors have been used to systematically
create gene knockouts in a variety of human cancer and immortalized cell
lines, in an effort to identify whether these knockouts cause cellular fitness
defects. Previous work has shown that CRISPR screens are more sensitive
and specific than pooled-library shRNA screens in similar assays, but
currently there exists significant variability across CRISPR library designs
and experimental protocols. In this study, we reanalyze 17 genome-scale
knockout screens in human cell lines from three research groups, using three
different genome-scale gRNA libraries. Using the Bayesian Analysis of Gene
Essentiality algorithm to identify essential genes, we refine and expand our
previously defined set of human core essential genes from 360 to 684 genes.
We use this expanded set of reference core essential genes, CEG2, plus
empirical data from six CRISPR knockout screens to guide the design of a
sequence-optimized gRNA library, the Toronto KnockOut version 3.0 (TKOv3)
library. We then demonstrate the high effectiveness of the library relative to
reference sets of essential and nonessential genes, as well as other screens
using similar approaches. The optimized TKOv3 library, combined with the
CEG2 reference set, provide an efficient, highly optimized platform
for performing and assessing gene knockout screens in human cell lines.

15
 INVESTIG ATION

A High-Density Linkage Map Reveals Sexual

Dimorphism in Recombination Landscapes in
Red Deer (Cervus elaphus)
Susan E. Johnston, Jisca Huisman, Philip A. Ellis, and Josephine M. Pemberton
G3: Genes | Genomes | Genetics August 2017, 7: 2859-2870

EDITORS’ NOTE Johnston et al. present a high-density linkage map for

a wild population of red deer (Cervus elaphus). Their investigation of the
recombination landscape shows a marked difference in recombination rates
between the sexes in proximity to the centromere, with females showing an
unusually elevated rate relative to other mammal species. This effect is most
pronounced in chromosomes that would have produced a new centromere
in the deer lineage. The authors propose that the observed effects may
have evolved to counteract selfish genetic elements that are associated with
asymmetrical female meiosis.

ABSTRACT High-density linkage maps are an important tool to gain insight

into the genetic architecture of traits of evolutionary and economic interest,
and provide a resource to characterize variation in recombination landscapes.
Here, we used information from the cattle genome and the 50 K Cervine
Illumina BeadChip to inform and refine a high-density linkage map in a wild
population of red deer (Cervus elaphus). We constructed a predicted linkage
map of 38,038 SNPs and a skeleton map of 10,835 SNPs across 34 linkage
groups. We identified several chromosomal rearrangements in the deer
lineage relative to sheep and cattle, including six chromosome fissions, one
fusion, and two large inversions. Otherwise, our findings showed strong
concordance with map orders in the cattle genome. The sex-averaged linkage
map length was 2739.7 cM and the genome-wide autosomal recombination
rate was 1.04 cM/Mb. The female autosomal map length was 1.21 longer than
that of males (2767.4 cM vs. 2280.8 cM, respectively). Sex differences in map
length were driven by high female recombination rates in peri-centromeric
regions, a pattern that is unusual relative to other mammal species. This
effect was more pronounced in fission chromosomes that would have had
to produce new centromeres. We propose two hypotheses to explain this
effect: (1) that this mechanism may have evolved to counteract centromeric
drive associated with meiotic asymmetry in oocyte production; and/or (2) that
sequence and structural characteristics suppressing recombination in close
proximity to the centromere may not have evolved at neo-centromeres.
Our study provides insight into how recombination landscapes vary and
evolve in mammals, and will provide a valuable resource for studies of
evolution, genetic improvement, and population management in red deer
and related species.

16
 INVESTIG ATION

High-Resolution Maps of
Mouse Reference Populations
Petr Simecek, Jiri Forejt, Robert W. Williams, Toshihiko Shiroishi, Toyoyuki Takada,
Lu Lu, Thomas E. Johnson, Beth Bennett, Christian F. Deschepper,
Marie-Pier Scott-Boyer, Fernando Pardo-Manuel de Villena, and Gary A. Churchill
G3: Genes | Genomes | Genetics October 2017, 7: 3427–3434

EDITORS’ NOTE Simecek et al. explore the genetic landscape of six mouse
reference populations by using a high-density genotyping array. Although the
strains were presumed to be fully inbred, they report residual heterozygosity
in 40% of individual mice from five of the six panels. The increased precision
offered by the new genetic maps let them identify de novo deletions
and duplications that ranged in size from 21 kb to 8.4 Mb in either the
homozygous or heterozygous state—as well as gene conversions.

ABSTRACT Genetic reference panels are widely used to map complex,

quantitative traits in model organisms. We have generated new high-
resolution genetic maps of 259 mouse inbred strains from recombinant inbred
strain panels (C57BL/6J × DBA/2J, ILS/IbgTejJ × ISS/IbgTejJ, and C57BL/6J
× A/J) and chromosome substitution strain panels (C57BL/6J-Chr#<A/J>,
C57BL/6J-Chr#<PWD/Ph>, and C57BL/6J-Chr#<MSM/Ms>). We genotyped
all samples using the Affymetrix Mouse Diversity Array with an average
intermarker spacing of 4.3 kb. The new genetic maps provide increased
precision in the localization of recombination breakpoints compared to the
previous maps. Although the strains were presumed to be fully inbred, we
found residual heterozygosity in 40% of individual mice from five of the six
panels. We also identified de novo deletions and duplications, in homozygous
or heterozygous state, ranging in size from 21 kb to 8.4 Mb. Almost two-thirds
(46 out of 76) of these deletions overlap exons of protein coding genes and
may have phenotypic consequences. Twenty-nine putative gene conversions
were identified in the chromosome substitution strains. We find that gene
conversions are more likely to occur in regions where the homologous
chromosomes are more similar. The raw genotyping data and genetic maps
of these strain panels are available at http://churchill-lab.jax.org/website/MDA.

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 INVESTIG ATION

Intricate and Cell Type-Specific Populations

of Endogenous Circular DNA (eccDNA) in
Caenorhabditis elegans and Homo sapiens
Massa J. Shoura, Idan Gabdank, Loren Hansen, Jason Merker, Jason Gotlib,
Stephen D. Levene, and Andrew Z. Fire
G3: Genes | Genomes | Genetics October 2017, 7: 3295–3303

EDITORS’ NOTE Shoura et al. highlight an understudied, mysterious

fraction of the eukaryotic genomes: endogenous circular DNA, which
are small circles of DNA found among the linear chromosomes. Using
an unbiased genome-wide approach, they provide an emerging
understanding of how fluctuations in genome 3D organization, as a
whole, contribute to cell fate and function.

ABSTRACT Investigations aimed at defining the 3D configuration of

eukaryotic chromosomes have consistently encountered an endogenous
population of chromosome-derived circular genomic DNA, referred to as
extrachromosomal circular DNA (eccDNA). While the production, distribution,
and activities of eccDNAs remain understudied, eccDNA formation from
specific regions of the linear genome has profound consequences on the
regulatory and coding capabilities for these regions. Here, we define eccDNA
distributions in Caenorhabditis elegans and in three human cell types,
utilizing a set of DNA topology-dependent approaches for enrichment and
characterization. The use of parallel biophysical, enzymatic, and informatic
approaches provides a comprehensive profiling of eccDNA robust to isolation
and analysis methodology. Results in human and nematode systems
provide quantitative analysis of the eccDNA loci at both unique and repetitive
regions. Our studies converge on and support a consistent picture, in which
endogenous genomic DNA circles are present in normal physiological states,
and in which the circles come from both coding and noncoding genomic
regions. Prominent among the coding regions generating DNA circles are
several genes known to produce a diversity

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 COLD FUSION Bullhead notothen, a red-blooded Antarctic fish, has just
11 chromosomes in its haploid set, but its recent ancestors had 24. A RAD-tag
meiotic map for the bullhead, along with comparative genomic analysis, showed
that massive karyotype reduction resulted from pairwise fusion of the 22 largest
chromosomes, followed by the two smallest chromosomes separately joining two
already fused chromosomes. Remarkably, no inversions mixed any of the fused
chromosome segments. The skull of this specimen was cleaned by Southern
Ocean isopods, whose voracious appetites make them the “dermestid beetles”
of the Southern Ocean. Photo: John H. Postlethwait.

Cold Fusion: Massive Karyotype Evolution in

the Antarctic Bullhead Notothen Notothenia coriiceps
Angel Amores, Catherine A. Wilson, Corey A. H. Allard,
H. William Detrich, and John H. Postlethwait
G3: Genes | Genomes | Genetics July 2017, 7: 2195–2207 19
 INVESTIG ATION

iSeq: A New Double-Barcode Method

for Detecting Dynamic Genetic
Interactions in Yeast
Mia Jaffe, Gavin Sherlock, and Sasha F. Levy
G3: Genes | Genomes | Genetics January 2017, 7: 143–153

EDITORS’ NOTE Systematic screens of genetic interactions have been used

in Saccharomyces cerevisiae to infer gene function, define network topology,
and study robustness; however, most screens characterize interactions in
a single environment. Jaffe, Sherlock, and Levy introduce iSeq—a double
barcoding platform that allows for rapid assessment of genetic interactions
across environments. They use iSeq to reproducibly measure fitness across
400 clonal strains of yeast and investigate the sources of variability in fitness
across replicate strains.

ABSTRACT Systematic screens for genetic interactions are a cornerstone of

both network and systems biology. However, most screens have been limited
to characterizing interaction networks in a single environment. Moving beyond
this static view of the cell requires a major technological advance to increase
the throughput and ease of replication in these assays. Here, we introduce
iSeq—a platform to build large double barcode libraries and rapidly assay
genetic interactions across environments. We use iSeq in yeast to measure
fitness in three conditions of nearly 400 clonal strains, representing 45
possible single or double gene deletions, including multiple replicate strains
per genotype. We show that iSeq fitness and interaction scores are highly
reproducible for the same clonal strain across replicate cultures. However,
consistent with previous work, we find that replicates with the same putative
genotype have highly variable genetic interaction scores. By whole-genome
sequencing 102 of our strains, we find that segregating variation and de novo
mutations, including aneuploidy, occur frequently during strain construction,
and can have large effects on genetic interaction scores. Additionally, we
uncover several new environment-dependent genetic interactions, suggesting
that barcode-based genetic interaction assays have the potential to
significantly expand our knowledge of genetic interaction networks.

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 INVESTIG ATION

Manipulating the Mitochondrial Genome

To Enhance Cattle Embryo Development
Kanokwan Srirattana and Justin C. St. John
G3: Genes | Genomes | Genetics July 2017, 7: 2065–2080

EDITORS’ NOTE Somatic cell nuclear transfer (SCNT) combines a donor cell
with a recipient oocyte to produce an embryo; this tool is commonly used in
cattle breeding, but the mixing of mitochondrial DNA (mtDNA) from the donor
and recipient can negatively impact embryo development. To combat this
problem, Srirattana and St. John used mtDNA-depleted cells as donors to
produce SCNT embryos that possess recipient oocyte-only mtDNA, along
with a reprogramming agent. Their results suggest this may become a useful
approach to enhancing livestock production traits.

ABSTRACT The mixing of mitochondrial DNA (mtDNA) from the donor cell
and the recipient oocyte in embryos and offspring derived from somatic
cell nuclear transfer (SCNT) compromises genetic integrity and affects
embryo development. We set out to generate SCNT embryos that inherited
their mtDNA from the recipient oocyte only, as is the case following natural
conception. While SCNT blastocysts produced from Holstein (Bos taurus)
fibroblasts were depleted of their mtDNA, and oocytes derived from Angus
(Bos taurus) cattle possessed oocyte mtDNA only, the coexistence of donor
cell and oocyte mtDNA resulted in blastocysts derived from nondepleted cells.
Moreover, the use of the reprogramming agent, Trichostatin A (TSA), further
improved the development of embryos derived from depleted cells. RNA-seq
analysis highlighted 35 differentially expressed genes from the comparison
between blastocysts generated from nondepleted cells and blastocysts
from depleted cells, both in the presence of TSA. The only differences
between these two sets of embryos were the presence of donor cell mtDNA,
and a significantly higher mtDNA copy number for embryos derived from
nondepleted cells. Furthermore, the use of TSA on embryos derived from
depleted cells positively modulated the expression of CLDN8, TMEM38A, and
FREM1, which affect embryonic development. In conclusion, SCNT embryos
produced by mtDNA depleted donor cells have the same potential to develop
to the blastocyst stage without the presumed damaging effect resulting from
the mixture of donor and recipient mtDNA.

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 INVESTIG ATION

Mechanisms of Transmission Ratio Distortion

at Hybrid Sterility Loci Within and Between
Mimulus Species
Rachel E. Kerwin and Andrea L. Sweigart
G3: Genes | Genomes | Genetics November 2017, 7: 3719–3730

EDITORS’ NOTE The hybrid offspring of recently diverged species often

show transmission ratio distortion, but the underlying genetic mechanisms
and evolutionary significance of this phenomenon are not always clear. In this
study, Kerwin and Sweigart identify multiple contributors to transmission ratio
bias in genomic regions that are linked to two Mimulus hybrid sterility loci.
They find little evidence of transmission bias within species, which suggests
that it might be limited to hybrid genetic backgrounds and not driven by
selfish evolution.

ABSTRACT Hybrid incompatibilities are a common correlate of genomic

divergence and a potentially important contributor to reproductive isolation.
However, we do not yet have a detailed understanding of how hybrid
incompatibility loci function and evolve within their native species, or why they
are dysfunctional in hybrids. Here, we explore these issues for a well-studied,
two-locus hybrid incompatibility between hybrid male sterility 1 (hms1) and
hybrid male sterility 2 (hms2) in the closely related yellow monkeyflower
species Mimulus guttatus and M. nasutus. By performing reciprocal
backcrosses with introgression lines (ILs), we find evidence for gametic
expression of the hms1-hms2 incompatibility. Surprisingly, however, hybrid
transmission ratios at hms1 do not reflect this incompatibility, suggesting
that additional mechanisms counteract the effects of gametic sterility.
Indeed, our backcross experiment shows hybrid transmission bias toward M.
guttatus through both pollen and ovules, an effect that is particularly strong
when hms2 is homozygous for M. nasutus alleles. In contrast, we find little
evidence for hms1 transmission bias in crosses within M. guttatus, providing
no indication of selfish evolution at this locus. Although we do not yet have
sufficient genetic resolution to determine if hybrid sterility and transmission
ratio distortion (TRD) map to the same loci, our preliminary fine-mapping
uncovers a genetically independent hybrid lethality system involving at least
two loci linked to hms1. This fine-scale dissection of TRD at hms1 and hms2
provides insight into genomic differentiation between closely related Mimulus
species and reveals multiple mechanisms of hybrid dysfunction.

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 SUNFLOWER SEEDS Although diploid Helianthus interspecific crosses are
expected to produce only diploid progenies, a significant proportion of triploid
progeny were observed while crossing several wild diploid perennial Helianthus
maximiliani and H. nuttallii populations with cultivated sunflower. Liu et al.
discovered that sunflower significantly increases triploid progeny production
through reduced incompatibility, which consequently increases the preferential
fertilization of pollen with slightly increased unreduced male gametes.
Photo: Zhao Liu, Gerald Seiler, and Chao-Chien Jan.

Triploid Production from Interspecific Crosses of Two

Diploid Perennial Helianthus with Diploid Cultivated
Sunflower (Helianthus annuus L.)
Zhao Liu, Gerald J. Seiler, Thomas J. Gulya, Jiuhuan Feng,
Khalid Y. Rashid, Xiwen Cai, and Chao-Chien Jan
G3: Genes | Genomes | Genetics April 2017, 7: 1097–1108
23
 INVESTIG ATION

TheCellMap.org: A Web-Accessible Database

for Visualizing and Mining the Global Yeast
Genetic Interaction Network
Matej Usaj, Yizhao Tan, Wen Wang, Benjamin VanderSluis, Albert Zou,
Chad L. Myers, Michael Costanzo, Brenda Andrews and Charles Boone
G3: Genes | Genomes | Genetics May 2017, 7: 1539–1549

EDITORS’ NOTE Usaj et al. present TheCellMap.org—a web-accessible

database and visualization tool designed to facilitate access to and
navigation of the global yeast genetic interaction network.

ABSTRACT Providing access to quantitative genomic data is key to

ensure large-scale data validation and promote new discoveries.
TheCellMap.org serves as a central repository for storing and analyzing
quantitative genetic interaction data produced by genome-scale Synthetic
Genetic Array (SGA) experiments with the budding yeast Saccharomyces
cerevisiae. In particular, TheCellMap.org allows users to easily access,
visualize, explore, and functionally annotate genetic interactions, or to
extract and reorganize subnetworks, using data-driven network layouts
in an intuitive and interactive manner.

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 MULTIPARENTAL POPUL ATIONS

Imputation-Based Fine-Mapping Suggests

That Most QTL in an Outbred Chicken
Advanced Intercross Body Weight Line Are
Due to Multiple, Linked Loci
Monika Brandt, Muhammad Ahsan, Christa F. Honaker, Paul B. Siegel,
and Örjan Carlborg
G3: Genes | Genomes | Genetics January 2017, 7: 119–128

EDITORS’ NOTE The Virginia chicken lines were founded over 60 years
ago to study the long-term effect of selection for two different phenotypes:
low- or high-body weight at 56 days of age. After 40 generations of such
selection, the lines displayed a nine-fold difference in body weight. Birds
from the high- and low-selected lines were crossed to found an Advanced
Intercross Line, which has been maintained for nine generations. Using
high-density genotypes of the founders, Brandt et al. imputed genotypes
in these intercross birds, which were only genotyped for a sparse set of
markers. Using single and multi-marker association analyses, they were able
to replicate nine body weight QTL.

ABSTRACT The Virginia chicken lines have been divergently selected for
juvenile body weight for more than 50 generations. Today, the high- and
low-weight lines show a >12-fold difference for the selected trait, 56-d
body weight. These lines provide unique opportunities to study the genetic
architecture of long-term, single-trait selection. Previously, several quantitative
trait loci (QTL) contributing to weight differences between the lines were
mapped in an F2-cross between them, and these were later replicated and
fine-mapped in a nine-generation advanced intercross of them. Here, we
explore the possibility to further increase the fine-mapping resolution of these
QTL via a pedigree-based imputation strategy that aims to better capture
the genetic diversity in the divergently selected, but outbred, founder lines.
The founders of the intercross were high-density genotyped, and then
pedigree-based imputation was used to assign genotypes throughout the
pedigree. Imputation increased the marker density 20-fold in the selected
QTL, providing 6911 markers for the subsequent analysis. Both single-marker
association and multi-marker backward-elimination analyses were used to
explore regions associated with 56-d body weight. The approach revealed
several statistically and population structure independent associations and
increased the mapping resolution. Further, most QTL were also found to
contain multiple independent associations to markers that were not fixed in
the founder populations, implying a complex underlying architecture due to
the combined effects of multiple, linked loci perhaps located on independent
haplotypes that still segregate in the selected lines.

25