Supplementary Data
Comparison of in vitro embryo and piglet editing frequency
For RELA TALEN mRNA injected zygotes we transferred 393 embryos into 11 recipient sows, resulting
in 6 pregnancies and 5 farrowings with litter sizes ranging from 3 to 17. One female aborted within
the last two weeks of pregnancy. Of the 46 piglets born, 5 were stillborn while of the live born 13
were savaged, sat on by the sow or culled on veterinary advice. Post mortem investigation failed to
detect any common pathology associated with the dead piglets.
In comparison to previous studies describing the combination of genome editing tools and SCNT,
zygote injection represents an efficient technology. In our work described here 32 donor/recipient
animals were used to produce 9 edited piglets. In contrast the generation of genome edited pigs by
SCNT used, for example, approximately 75 donor/recipients to produce 2 edited animals12 while
approximately 84 donor/recipient animals were used to produce 11 edited piglets in a report of
biallelic genome editing10; assuming 20 oocytes per donor animal.
Supplementary Fig 1. Example TALEN edited pig (#26) carrying single base deletion with sequence
read of single base pair edited allele and wild-type RELA allele.
TALEN binding site
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
Zygote 1
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCC~~~~TGATGGAGTACCCTGAGGCTATAACTC 4
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTAGTGGAGTACCCTGAGGCTATAACTC 2+2
Zygote 2
GGTGTATCCATGCCCCCCCACACAGCTGAG~CCATGCTGATGGAGTACCCTGAGGCTATAACTC 1
Zygote 3
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~~~~~ATGGAGTACCCTGAGGCTATAACTC 5
Zygote 4
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATTGCTGATGGAGTACCCTGAGGCTATAACT +1
Zygote 5
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCC~TGCTGATGGAGTACCCTGAGGCTATAACTC 1
Zygote 6
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCC~~~~TGATGGAGTACCCTGAGGCTATAACTC 4
Zygote 7
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~GCTGATGGAGTACCCTGAGGCTATAACTC 1
Zygote 8
GGTGTATCCATGCCCCCCCACACAGCTGAGCC35TGCTGATGGAGTACCCTGAGGCTATAACTC 2+35
ACAAGTCGAGGGTCTCAGAGTCACTGAGTACTGAG
Zygote 9
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTACCCATGCCCCCCCACACAGCTGAGCCCA~~~~~ATGGAGTACCCTGAGGCTATAACTC 5
Zygote 10
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCT~~~~GATGGAGTACCCTGAGGCTATAACTC 4
Zygote 11
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCC~ATGCTGATGGAGTACCCTGAGGCTATAACTC 1
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGG~~~~CCCTGAGGCTATAACTC 4
Zygote 12
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCC~ATGCTGATGGAGTACCCTGAGGCTATAACTC 1
Zygote 13
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATG~TGATGGAGTACCCTGAGGCTATAACTC 1
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCAT~~TGATGGAGTACCCTGAGGCTATAACTC 2
Zygote 14
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCC~ATGCTGATGGAGTACCCTGAGGCTATAACTC 1
Zygote 15
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCC~~~~TGATGGAGTACCCTGAGGCTATAACTC 4
Zygote 16
GGTGTATCCATGCCCCCCCACACAGCTGAGC~~ACACA~~~GGAGTACCCTGAGGCTATAACTC 10+5
Zygote 17
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCT~~~~~~~~~~~GATGGAGTACCCTGAGGCTATAACTC 11
Zygote 18
GGTGTATCCATGCCCCCCCACACAGCT~~~~~~~~~~~~~~~GAGTACCCTGAGGCTATAACTC 15
Zygote 19
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~~~TGATGGAGTACCCTGAGGCTATAACTC 3
Zygote 20
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGA~~~~~~GCTGATGGAGTACCCTGAGGCTATAACTC 6
Zygote 21
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCACAGCTGGTACTCAGATGGAGTACCCTGAGGCTATAACT +9
GGTGTATCCATGCCCCCCCACACAGCT~~~~~~~~~~~GATGGAGTACCCTGAGGCTATAACTC
11
Supplementary Fig 2. Sequence of indels identified in zygotes. Zygotes 1-16 were injected with
20ng/ul TALEN mRNA and screened for GFP. Zygotes 17 & 18 were injected with 20ng/ul TALEN
mRNA and were screened for development. Zygotes 19 – 21 were injected with 2ng/ul TALEN mRNA.
TALEN binding site
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
Piglet 8770-I
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~GCTGATGGAGTACCCTGAGGCTATAACTC 1
Piglet 8770-J
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATTGCTGATGGAGTACCCTGAGGCTATAACT +1
Piglet 8770-26
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~GCTGATGGAGTACCCTGAGGCTATAACTC 1
Piglet 8130-so
GGTGTATCCATGCCCCCCCACACAGCTGA~~~~~~GCTGATGGAGTACCCTGAGGCTATAACTC 6
Piglet 8130-16
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCATGCTGATGGAGTACCCTGAGGCTATAACTC WT
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCTCCATCAGCTGATGGAGTACCCTGAGGCTAT +5
GGTGTATCCATGCCCCCCCACACAGCTGAG~~~~~~~~~~~~~~~~~CCCTGAGGCTATAACTC 17
Inserted sequence is duplication/inversion of orange highlighted
Piglet 8784-30
GGTGTATCCATGCCCCCCCACACAGCTGAGC~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ 111
GGTGTATCCATGCCCCCCCACACAGCTGAGCC~~~~CTGATGGAGTACCCTGAGGCTATAACTC 4
Piglet 8784-33
GGTGTATCCATGCCCCCCCACACAGCTGAGC~~~~~CTGATGGAGTACCCTGAGGCTATAACTC 5
GGTGTATCCATGCCCCCCCACACAGCTGAG~~~~~~CTGATGGAGTACCCTGAGGCTATAACTC 6
GGTGTATCCATGCCCCCCCACACAGCTGAGCC~~~~CTGATGGAGTACCCTGAGGCTATAACTC 4
Piglet 8784-34
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCA~GCTGATGGAGTACCCTGAGGCTATAACTC 1
GGTGTATCCATGCCCCCCCACACAGCTGAGCCCTGAGTACCCTGAGGAGTACCCTGAGGCTATA 8+12
Inserted sequence is duplication of orange highlighted
ZFN binding site
TGCTGATGGAGTACCCTGAGGCTATAACTCGCTTGGTGACAGGGTCCCAGAGACCCCCTGACC WT
Piglet 8142-C
TGCTGATGGAGTACCCTGAGGCTATAACTC~~~~~~~~~TCTGGGA~~~~~~~~~CCCTGACC 25+7
Inserted sequence is inversion of orange highlighted in consensus
Supplementary Fig 3. Sequence of indels identified in piglets.