Genetic architecture of developmental traits in

populations of male gypsy mothsChristopher J. Friedline, Ph.D.!

Virginia Commonwealth University

@noituloveand @cfriedline

Evolution 2014!Raleigh, NC!6.21.2014

Lymantria dispar

http://bugguide.net/node/view/553307Adapted from: http://www.fs.fed.us/ne/morgantown/4557/gmoth/atlas/quar1a.gif

http://green.blogs.nytimes.com/2011/09/13/the-toll-from-tree-boring-pests/?_php=true&_type=blogs&_r=0

Can we gain insight into the genetic architecture of invasion

using L. dispar as a model?

Experimental design• 7 populations established in

replicated common gardens (NY, VA)

• NC (1), VA (2), NY (1), Quebec (2) + 1 lab strain (CPHST Otis Lab)

• Single reference assembly (HiSeq 2500 PE)

• 188 barcoded individuals

• 3 phenotypes (developmental)http://www.fs.fed.us/ne/morgantown/4557/gmoth/

Parry and Grayson, EntSoc 2013

Reference Assembly• MaSuRCA Zimin et al. (2013), Bioinformatics, 29 (21),

2669-2677

• 591,450 scaffolds (90M QC PE reads)

• 588,164,124 Mb (~1 Kb/contig, max=28 Kb)

• 268 contigs > 10 Kb

• N50 = 1.3 Kb

• 35.3% GC

• 32%/56% CEGMA complete/partial

ddRADseq

• 316 M reads (41.5 GB)

• 188 individuals

• 690 Kb/individual [10, 1400] Kb (QC)

Peterson et al. (2012) Plos One 7.5 (2012): e37135-e37135

SNP Calling/Filtering• Called with Freebayes (n = 30,791)

• Kept only biallelic SNPs

• Removed > 50% missing

• Removed FIS outliers (> |0.5|)

• Removed MAF < 0.01

• n = 11,021

Phenotypes

NC NY OTIS QC32 QC93 VA1 VA2

7

8

9

10

11

12

13

Pupual Duration

NC NY OTIS QC32 QC93 VA1 VA2

0.2

0.3

0.4

0.5

0.6

0.7Mass

NC NY OTIS QC32 QC93 VA1 VA2

65

70

75

80

85

90

95

Total Dev Time

0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35FST

0

1000

2000

3000

4000

5000

6000

7000

8000

Multilocus FIS = 0.0486, FST = 0.0224, FIT = 0.0700

n=11021 [0.00, 0.34]

FST

MAF Bin F F F

50 0.1542 0.0234 0.174

40 0.1007 0.0303 0.128

30 0.0757 0.0295 0.103

20 0.0093 0.0169 0.0261

10 -0.0673 0.0088 -0.0579

Population Structure Correction by PCA

• Price et al. (2006) ”Principal Components Analysis Corrects for Stratification in Genome-wide Association Studies." Nat Genet 38.8

• First principal component approximates FST

• Number of axes chosen using a Tracy-Widom test, described in Eckert et al. (2010). Genetics 185: 969-982.

• Correlation of genotype vs. phenotype residuals to X2 to p-values

Population Structure Correction by PCA

• Price et al. (2006) ”Principal Components Analysis Corrects for Stratification in Genome-wide Association Studies." Nat Genet 38.8

• First principal component approximates FST

• Number of axes chosen using a Tracy-Widom test, described in Eckert et al. (2010). Genetics 185: 969-982.

• Correlation of genotype vs. phenotype residuals to X2 to p-values

�40 �20 0 20 40 60 80

PC1 (0.021%)

�60

�50

�40

�30

�20

�10

0

10

20

30

PC2

(0.0

15%

)

PCA of n=7 populations on 11021 loci

Population Structure Correction by PCA

• Price et al. (2006) ”Principal Components Analysis Corrects for Stratification in Genome-wide Association Studies." Nat Genet 38.8

• First principal component approximates FST

• Number of axes chosen using a Tracy-Widom test, described in Eckert et al. (2010). Genetics 185: 969-982.

• Correlation of genotype vs. phenotype residuals to X2 to p-values

Top SNP

C/C C/T T/TLocus 14103 (ctg7180001511349/152)

0.2

0.3

0.4

0.5

0.6

0.7Mass (p = 0.000004, FST = �0.008543)

T/T T/C C/CLocus 14908 (ctg7180001527347/31)

65

70

75

80

85

90

95

Total Dev Time (p = 0.000111, FST = 0.016849)

A/A A/T T/TLocus 10529 (ctg7180001452692/364)

7

8

9

10

11

12

13

Pupual Duration (p = 0.000022, FST = �0.024810)

468444

44

444

73 29

9

Total Dev Time Pupual Duration

Mass

• Corrected for population structure (Price et al. 2006), binned by MAF

• By p value (p < 0.05):

• Mass: n = 555

• Pupual duration: n = 526

• Total development time: n = 524

• By q value (Storey and Tibshirani, 2003)

• Mass: n = 3 (14103(*,10), 27843(40), 9023(40))

• Pupual duration: n = 1 (S10529(40))

• Total development time: n = 0

Significant SNPs

Multilocus effectS2

7843

S533

0S1

3360

S230

45S2

3046

S146

81S1

4682

S278

41S3

0469

S258

51S1

0170

S237

91S1

2194

S259

17S7

263

S787

0S1

134

S141

03S8

633

S132

28S1

4762

S260

9S2

6153

S178

55S9

023

Top 25 (p-value) loci

�0.005

0.000

0.005

0.010

mul

t.re

g.co

eff

Mass regression coefficients(R2 = 0.556, R2

adj = 0.494)

S100

56S2

0804

S305

88S2

1213

S114

90S2

3793

S298

19S1

8637

S140

30S2

1212

S125

63S2

4930

S261

20S2

3756

S238

92S1

2194

S149

08S1

5702

S122

18S1

5492

S301

68S2

9721

S287

52S1

9836

S100

57

Top 25 (p-value) loci

�1.0

�0.5

0.0

0.5

1.0

mul

t.re

g.co

eff

Total Dev Time regression coefficients(R2 = 0.639, R2

adj = 0.584)

S160

63S6

315

S101

93S1

8784

S218

58S5

008

S215

6S1

8532

S184

65S1

2721

S120

12S2

8904

S257

1S2

0272

S696

3S3

0060

S319

6S3

195

S165

60S1

7662

S105

29S2

0271

S185

30S6

316

S160

62

Top 25 (p-value) loci

�1.0

�0.5

0.0

0.5

1.0

1.5

mul

t.re

g.co

eff

Pupual Duration regression coefficients(R2 = 0.541, R2

adj = 0.473)

Blast resultsMass

reverse transcriptase*

non-LTR retrotransposon

predicted craniofacial development protein

sulfotransferase (amine, estrogen)

Pupual duration

endonuclease-reverse transcriptase

phosphatidylinositol 3-kinase

Development time

reverse transcriptase

endonuclease-reverse transcriptase

transcription initiation factor TFIID subunit 2-like protein

chosen by p+q*!chosen by q+

Conclusions/Future Work• Assembly curation is likely necessary for more

robust biological conclusion

• Small effect sizes difficult to detect with small sample size and populations

• High degree of multilocus effects

• Additional replicate gardens with related material

• Probabilistic genotype calling with full set

Acknowledgments

Johnson Lab!Derek Johnson

Kristine Grayson Trevor Faske

NPGI: NSF Postdoctoral Fellowship in Biology FY 2013

Rodney Dyer, VCU Dylan Parry, SUNY-ESF

Eckert Lab!Andrew Eckert Brandon Lind Erin Hobson

Ethan Harwood VCU NARF VCU CHiPC