JULY 2017

Rewriting DNA Synthesis

Emily Leproust, PhD

CEO

@TwistBioscience #SyntheticDNA

Founded

2013

San Francisco

South San Francisco

Tel Aviv

Raised $195M

Funding

2

3

Delivering Game-Changing

Throughput, Speed, Cost and

Quality

121 Oligos

Many New Applications for Synthetic DNA

AUGUST 7 , 2017 TW IST CONFIDENTIAL 4

Need a New Typeof DNA Supplier to

Meet Demand

Need a New Typeof DNA Supplier to

Meet Demand

CURRENT DNATECHNOLOGIES

RecombinantDNA

GenomicAdvances

Cloning

AgriculturalOil-Free Fertilizers

Drought Solutions

New Disease Protection

Biotech / PharmaAntibodies / TCR

Vaccines

Immuno and Cancer Therapies

Small Molecule Drug Mfg.

Industrial Bio-ProductionSpecialty Chemicals

Advanced Property Materials

Today’s Market: Buyers and Makers

5AUGUST 7 , 2017 TW IST CONFIDENTIAL

Small Scale, Academic Users

Price-Sensitive

Large Scale, Commercial Users

Value Speed, Throughput and

Quality

$1B / Year

“Buy”

Synthetic DNA

$250M

“Make”Synthesis Supplies

(Enzymes, Primers, Cells,

Plates)$750M

Can’t GetWhat I Need!

“ ”I Hate Cloning!

“ ”

Source: Percepta Associates

Current Technologies Are Limiting

AUGUST 7 , 2017 TW IST CONFIDENTIAL 6

PROCESS BOTTLENECK

• Too long

• Can’t scale up

or down

• Expensive

Select/Screen for Desired FunctionE.g., Bacteria, Yeast, Algae Bio-Factories, Plants,

Mammalian Cells

Design Gene Sequences1 to 10,000+ Variants Possible per Cycle

AGCTGCTCGAATACGATAG

Manufacture SyntheticGene Constructs• Genes (1k Basepairs)

• Parts (3 - 10 Genes)

• Pathways (10 - 100 Genes)

• Chassis (100 - 1000 Genes)

Build

Test

Cycles5-10x

Want 1 Gene in 2 Weeks $0.2- $1/bp or Non-Clonal + Work

Want 10,000 Genes in 2 Weeks Use 3 Vendors and/or Wait 3 Months

Limitations of 20-Year Old Technology

7

96 Well Plastic Plate

96 Oligos = 1 Gene

Does Not Scale to Meet Market

Need

Does Not Scale to Meet Market Need

Rewriting DNA Synthesis with the

Power of Silicon

AUGUST 7 , 2017 TW IST CONFIDENTIAL 8

Developing Game-Changing Throughput, Speed, Cost and Quality

96 Oligos 1 Gene

TRADIT IONAL METHODS

Based on 96-well plateTWIST BIOSCIENCE

Silicon-based Synthesis Platform

> 1 Million Oligos 9,600 Genes

3.2kb genes are at the core of various pipelines

Antibody-based

drug development

Up to 3.2 kb

Gene editing:

donor DNA synthesis

Photo sources:

medicalxpress.com, plos.org

Pathway assemblies

Gene therapy

10

Your Sequence,

Your Way

• Twist’s or your vector

• 100% NGS Verified

• up to 3.2kb

Industry Leading

Price & Fast

Turnaround Time

• 9c/bp

• 15-20 days TAT

Platform Scalability

• No order limits

• 1 – 10,000+ genes

Why Clone? Let Twist Build It

G E N E S

11

Your Sequence,

Your Way

• >1 error 3,000 bp

Industry Leading

Price & Fast

Turnaround Time

• 7c/bp

• 7 days TAT

Platform Scalability

• No order limits

• 1 – 10,000+ genes

Need Linear DNA? Let Twist Build It

N O N - C L O N A L F R A G M E N T S

Market Leader in Gene Synthesis

12

G E N E S P R O D U C E D A N D S H I P P E D

0

5,000

10,000

15,000

20,000

25,000

30,000

35,000

40,000

Jan-Mar 16 Apr-Jun 16 Jul-Sep 16 Oct-Dec 16 Jan-Mar 17

Shipped

Produced

Apr – Jun 2016: 2,000 genes per month

Jan – Mar 2017: 10,000 genes per month

December 2017: 60,000 genes per month

We Can Make

1 Million

Guide RNA Pairs

D E S I G N & B U I L D

P A R T N E R S H I P

Fueling Genome-Wide Experiments

13

C R I S P R - C A S 9 S Y S T E M

Novel Protein Libraries for Drug DiscoveryEnabling Efficiency in Drug Development

14

From Needlein a Haystack

• Random diversity

• Biased representation

• >99% inefficiency

• Lengthy optimization

cycle

• Expensive process

Gene Synthesis

Single Site

Multi-Site

Stretch

Multi-Domain

Precise Introduction of Variants,

Diversity that Enables Screening

Efficiency

.

. .

.

.

.

.

.

. .

Gene Synthesis

Single Site

Multi-site

Stretch

Multi-Domain

...

.

..

To Stack of Needles

• Explicit

• Even representation

• Human repertoire

based

• Fast

• Affordable

Protein Drug Discovery

½ the time

¼ the cost

15

P R E C I S I O N L I B R A R I E S

High Quality Precision

Libraries

• Single Site Variant Libraries

• Combinatorial Libraries

High Diversity

• 104 to 1010 variants

• 100% explicit sequence,

no random mutations

Industry Leading Price &

Rapid Turnaround Time

• ½ the time

• ¼ the cost

vs.

codon-by-codon

synthesis

52

1

GFPPX

Assay Development

Construct

mutagenesis library

Pharmacological

Characterization

Circuit integration

DNA Assembly ScreeningTransformation

Rational Design vs Random Mutagenesis

CH

AR

AC

TE

RIZ

AT

IO

N

Sequencing

PX

PX

Acknowledgements:

David Oling

Lina Lawenius

Niklas Larsson

Mark Wigglesworth

Tom Ellis

William Shaw

4

Random mutagenesis

(epPCR)

VS.

Saturation mutagenesis

Twist Saturation Mutagenesis Library

Highly uniform representation of every mutant

across all 161 positions

Less Work, More Hits

CriteriaTwist

Saturation libraryError prone PCR library

Variants All variants present Unknown

Number of Hits 10/10 hits2/10 (both present in Twist

library)

Overall

+ More hits

+ Pure, cloning ready DNA

+ Fast production time

- Less hits

- Unknown variants

- Large fraction of empty vectors

Finely Tuned Precision Libraries

Objective: Introduce variants at low frequency to mimic natural

distribution within the human immune repertoire

*Sequence and data anonymized

SPRIPWLCI??F?QVAE?LL??FEAHDPHPAACLFLIIPGH

Variable Positions 1 2 3 4 5 6

Total Diversity 1,391,208

Highly Precise Variant Introduction

Position 1

Expected Observed

A 2.7% 2.8%

C 8.6% 8.0%

D 2.7% 2.7%

E 8.6% 8.4%

F

G 1.7% 1.5%

H 8.6% 8.8%

I

K

L 8.6% 8.9%

M 1.7% 1.5%

N 1.7% 1.8%

P 2.7% 2.8%

Q 8.6% 7.6%

R 8.6% 7.9%

S 8.6% 8.3%

T 8.6% 9.0%

V

W

Y

0.00%

1.00%

2.00%

3.00%

4.00%

5.00%

6.00%

7.00%

8.00%

9.00%

10.00%

A C D E F G H I K L M N P Q R S T V W Y

Position 1Expected

Observed

Χ2 = 1

*Sequence and data anonymized

Highly Precise Variant Introduction

Position 2

Expected Observed

A 2.7% 2.9%

C 6.3% 6.6%

D 2.7% 2.7%

E 6.3% 6.2%

F

G 7.7% 7.4%

H 6.3% 6.3%

I

K

L 6.3% 6.4%

M 7.7% 7.0%

N 7.7% 8.5%

P 2.7% 2.8%

Q 6.3% 5.4%

R 6.3% 5.9%

S 6.3% 5.9%

T 6.3% 6.2%

V

W

Y

0.00%

1.00%

2.00%

3.00%

4.00%

5.00%

6.00%

7.00%

8.00%

9.00%

A C D E F G H I K L M N P Q R S T V W Y

Position 2Expected

Observed

Χ2 = 1

*Sequence and data anonymized

Powering the Synthetic

Biology Revolution

Thank you!

• Appendix