Agi apr13 aylings

The Genome Analysis Centre

Building Excellence in Genomics and Computational Bioscience

Sarah AylingComputational Genomics Group Leader

[email protected]

Rice resequencing


The Genome Analysis CentreThe Genome Analysis CentreThe Genome Analysis Centre

Rice Resequencing Project

Aim: To bring genomics capability to rice breeding in Vietnam in light of changing climates

Approach: Sequence varieties with interesting phenotypes and provide training in bioinformatics

The Genome Analysis CentreThe Genome Analysis Centre

Training at TGAC

Two scientists from AGI visited TGAC in September 2012 for bioinformatics training

Training topics: NGS assembly and alignment tools Phylogenetics Browser training Variant calling



Varieties


Stress Varieties: indica, japonica and javanica

Bacterial Blight Resistance

Hom rau; Khau dien lu; Nep meo nuong; Tep Thai Binh; Toc lun

Blast Resistance Ble te lo; Khau mac Buoc; Chiem nho Bac Ninh 2; Nep lun; OM 6377

Brown Planthopper Resistance

Chan thom; Coi ba dat; Khau giang; OM5629; Xuong ga

Drought tolerance Ba cho K’te; Blao sinh sai; Tan ngan; Nang quot bien; Nep bo hong Hai Duong;

Quality potential Nang thom cho Dao; Tam xoan Bac Ninh; Tam xoan Hai Hau; Te Nuong; OM 3536; Thom lai

Salt tolerance Lua Ngoi; Mot bui do; Chiem do; Nang co do 2; Nep man

Unclassified Nep ong tao; Khau Lien; Lua goc do; Chiem da; IS1.2



Sequencing


Illumina HiSeq 2000

• DNA sheared into fragments 300-500 bp in length

• 100bp sequenced from both ends of fragments

• 18 lanes of sequencing (2.25 flowcells)

• 1.3 Tb of sequence data



Ch i

em d

a

Ch i

em d

o

Ch i

em n

ho B

ac N

inh

2

IS1 .

2

Lua

goc

do

Na n

g c o

do

2

Ne p

bo

hon g

Ha i

Du o

ng

Ne p

lun

Ne p

ma n

OM

353 6

OM

637 7

Te p

Th a

i Bi n

h

Th o

m L

ai

To c

lun

Xu o

ng g

a

Ba

cho

K’te

Bla

o si

n h s

a i

Ble

te lo

Ch a

n th

om

Co i

ba

dat

Ho m

ra u

Kh a

u d i

en lu

Kh a

u g i

ang

Kh a

u L i

en

Kh a

u m

ac b

uoc

Lua

Ng o

i

Mo t

bu i

do

Na n

g th

om c

ho d

ao

Ne p

me o

nu o

ng

Ne p

on g

tao

OM

562 9

Ta m

xo a

n B

ac N

inh

Ta m

xo a

n H

ai H

au

Ta n

ng a

n

Te

Nuo

ng

Na n

g q u

ot b

ien

0

10

20

30

40

50

60

70

Varieties

Co

ver a

ge

(a

ssu

mi n

g 4

30

Mb

ge

no

me

siz

e)

Sequencing depth


Average coverage Detect homozygous base p(0.9975)

30x Detect heterozygous base p(0.9975)



Reference genomes


Indica: 93-11 Yu et al., Science 2002

Temperate japonica: Nipponbare Goff et al., Science 2002



Align reads to references (BWA)


Ch i

em d

a

Ch i

em d

o

Ch i

em n

ho B

ac N

inh

2

IS1 .

2

Lua

goc

do

Na n

g c o

do

2

Ne p

bo

hon g

Ha i

Du o

ng

Ne p

lun

Ne p

ma n

OM

353 6

OM

637 7

Te p

Th a

i Bi n

h

Th o

m L

ai

To c

lun

Xu o

ng g

a

Ba

cho

K’te

Bla

o si

n h s

a i

Ble

te lo

Ch a

n th

om

Co i

ba

dat

Ho m

ra u

Kh a

u d i

en lu

Kh a

u g i

ang

Kh a

u L i

en

Kh a

u m

ac b

uoc

Lua

Ng o

i

Mo t

bu i

do

Na n

g th

om c

ho d

ao

Ne p

me o

nu o

ng

Ne p

on g

tao

OM

562 9

Ta m

xo a

n B

ac N

inh

Ta m

xo a

n H

ai H

au

Ta n

ng a

n

Te

Nuo

ng

Na n

g q u

ot b

ien

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

unalignedJaponica-specificIndica-specificaligned to both

Varieties

Per

cen t

age

of re

ads

indicas japonicas javanica



SNP discovery


Align reads to references using BWA v0.6.1

Detect variants using GATK v1.6

Insertion/deletion realignment

SNP calling and filtering



SNPs by reference


0 200000 400000 600000 800000 1000000 1200000 1400000 16000000

200000

400000

600000

800000

1000000

1200000

1400000

1600000

Indicas

Japonicas

javanica

Number of SNPs on indica reference

Num

ber

of S

NPs

on

japo

nica

ref

eren

ce



Heterozygosity


Average % heterozygous SNPs: 0.14%

Range % heterozygous SNPs: 0.05-0.38%

Average Ts/Tv: 2.39

Range Ts/Tv: 1.95-2.55



Grouped SNPs


Trait Number of shared SNPs

Unique to set

Shared with 1

Shared with 2

Shared with 3

Shared with 4

Shared with 5

Shared with 6

Blast 43,755 - - - - 2 6 6

Blight 22,065 - - 1 20 9 15 32

Drought 27,109 - - - - 1 2 3

Planthopper 43,238 - - - 1 - 7 20

Quality 55,256 - - - 1 8 14 30

Salt 51,472 - - 3 9 18 56 149


tgac-browser.tgac.ac.uk

Username: viet_ricePassword: v1et_r1ce


The Genome Analysis CentreThe Genome Analysis CentreThe Genome Analysis Centre

Acknowledgements

Mario CaccamoSarah Ayling

Melanie FebrerAnil Thanki

Xingdong Bian

Prof HamKhuat Huu Trung

Khoa Nguyen Truong and colleagues

Giles OldroydChristian Rogers


THANKS!

Technology

Agi apr13 aylings