1. Dept. of Plant Breeding & Genetics, Cornell University
Jahn Lab
Dept. of Plant Breeding & Genetics, Cornell University
Original Objectives Years 2-4
Generate a BAC library from Capsicum frutescens, a major source of disease resistance in pepper
Fine map the L locus in Capsicum and
assess homology to R3/R7/potato
and I2/tomato
3. Provide 3 BACs to TIGR for sequencing
Comparative structural analysis of
orthologous R gene clusters
Thank you for the kind introduction.
It is my great honor to have an opportunity to give a job talk today.
And I would like to express my thanks to search committee members and chairman of this department.
The title that I’m going to talk today is “Developing a platform for the future vegetable breeding: Capsicum as an example”.

2. Dept. of Plant Breeding & Genetics, Cornell University
Jahn Lab
Dept. of Plant Breeding & Genetics, Cornell University
Current Objectives Years 2-4
Generate a BAC library from Capsicum frutescens, a major source of disease resistance in pepper
Fine map the L locus in Capsicum and assess homology to R3/R7/potato and I2/tomato
Provide 10 Capsicum BACs to TIGR for draft sequencing, identify clones for full sequencing
Comparative structural analysis of
homologous NBS-LRR containing regions
Provide Capsicum-derived transgenes
for resistance to PVY for use in
potato transformation and breeding
Thank you for the kind introduction.
It is my great honor to have an opportunity to give a job talk today.
And I would like to express my thanks to search committee members and chairman of this department.
The title that I’m going to talk today is “Developing a platform for the future vegetable breeding: Capsicum as an example”.

3. Current Project Objectives
■ Generate a BAC library from Capsicum frutescens
■ Improve map of the L locus in Capsicum with clones from potato and tomato maps
■ Assess homology to R3/R7/potato and I2/tomato
■ Build Capsicum contig through selected region (I2)
■ Obtain draft sequence to identify BAC clones for full sequence; obtain full sequence
■ Comparative assessment of sequence from tomato, potato and pepper chromosome 11
■ Developing potyvirus resistant potato using Capsicum eIF4E sequences
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

4. Comparative study of R3/R7 loci
■ BAC library from Capsicum frutescens
Leaf tissues of C. frutescens # of clones: 221,184 clones average insert size: kb BAC library has already been widely used
in Jahn lab: for studies of the Pun1 locus
for studies of the L locus
in Paran lab: for studies of a number of loci involved with fruit color and fruit quality available at cost from AGI
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

5. Some cross-generic R gene groups in Solanaceae

6. Potato 11 Tomato 11 Pepper 11 > 30 cM 6.0 cM AFLP markers < 1.0
CT107
CT107
TG105, I2?
TG46
TG46
TG46
TG36
> 30 cM
TG36
6.0 cM
TG105, I2
TG105, I2
T142, CDI 136
R3 BAC contig R3
6H15
AFLP markers < 1.0 L
TG36
T1151
T386

7. Comparative mapping R3/R7 TG105 and I2 orthologs in pepper map 20 cM north of Capsicum L locus
potato 11
tomato 11
pepper 11
CT107
CT107
TG105, I2?
TG46
TG46
TG46
TG36
TG36
TG105, I2
TG105, I2
T142, CDI 136
BAC contig R3
6H15
AFLP markers < 1.0 L
TG36
T1151
T386

8. Overview of our strategy to characterize I2-homologous Capsicum regions
BAC library screening
I2, GP250
Positive BAC clone analysis
BAC fingerprinting and contig mapping
PCR screening using primers developed in potato
Candidate BAC clone draft sequencing
Map BAC sequences on a pepper linkage map
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.
Identify BAC clones orthologous to R3/R7
Complete sequencing orthologous clones

9. Cloning homologs of BAC end sequences

10. Identification of BAC clones
■ Probes I2C-1 gene: 89 positive pepper clones GP250: 7 positive pepper clones These sets did not overlap
There are 17 copies of GP250 and I2 homologs within
the R3/R7 cluster in potato.
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.
Implies significant differences in genome organization between
potato and pepper in this region

11. BAC fingerprinting and contig mapping
Clone 70
Clone 36
Clone 59, 92
Clone 81
Clone 82
Clone 47, 29
Clone 49
Clone 83
Clone 67
Clone 63
Clone 19
Clone 39
Clone 91
Clone 42, 85
Clone 75
Clone 68
Clone 41
Clone 56
Clone 61
Clone 1
Clone 40
Clone 48, 60
Clone 50
Clone 31, 79
Clone 30, 35, 44, 71
Clone 62
Clone 87
Clone 7
Clone 35, 43, 54, 73, 74, 78
Clone 32, 34
Clone 76
Clone 66
Clone 52, 84
Clone 57
Clone 86, 90
Clone 88, 89
Clone 80
Clone 33
Clone 38, 51, 69
Clone 37
Clone 77
Clone 45, 46, 58, 65
Clone 12
Clone 22
Clone 21
Clone 27
Clone 25
Clone 4
Clone 28
Clone 26
Clone 72
Clone 23
Clone 8, 9
Clone 10
Clone 14, 17
Clone 14, 15
Clone 2
Clone 13, 18
Clone 6
Clone 5, 11, 13, 20
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

12. Analysis of I2 positive pepper clones
■ PCR screening I2 positive BAC clones
Primers: R7-1, R7-2, LRR, BAC end sequences
R7-1: 37 BAC clones, 1.5 kb
R7-2: 52 BAC clones, 0.8 kb to 1.3 kb
LRR: 22 BAC clones, 1.2 kb
BAC end sequences: none
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.
■ Cloning PCR fragments and sequencing
The BAC clones obtained by LRR primers.

13. Sequence alignment of LRR fragments1 2 3 4 5 6 7 8 9
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean. 10
Ten BAC clones were chosen for draft sequencing
Sent to TIGR mid-January, 2006

14. Summary and Future Plans
■ A total of 89 positive BAC clones were identified by I Seven BAC clones positive to GP250 were also identified. None of GP250 clones cross hybridized with I2 positive clones. ■ Ten BAC clones were chosen for draft sequencing by BAC fingerprinting and PCR analysis. ■ Draft BAC sequences will be mapped on a pepper linkage map to find pepper sequences orthologous to R3/R7. ■ Complete sequences of the orthologous pepper BAC clones will be determined and compared with potato sequences
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

15. Summary and Future Plans
■ A total of 89 positive BAC clones were identified by I Seven BAC clones positive to GP250 were also identified. None of GP250 clones were cross hybridized with I2 positive clones. ■ Ten BAC clones were chosen for draft sequencing by BAC fingerprinting and PCR analysis. ■ Draft BAC sequences will be mapped on a pepper linkage map to find orthologous sequences to R3/R7. ■ Complete sequences of the orthologous BAC clones will be determined and compared with potato sequences
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

16. Project Goals
■ Comparative study of R3/R7 loci in potato and their orthologous in pepper
■ Developing potyvirus resistant potato using potyvirus resistance genes from pepper
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

17. Potato Virus Y ■ Potato virus Y ■ Strains
- Infects the Solanaceae crops: potato, tobacco, tomato and pepper
- One the most economically important plant pathogens
- Transmitted by aphids non-persistently
- Tuber-borne virus in potato
■ Strains
- PVY N: causes veinal necrosis and mild mottling on N. tabacum cv.
Xanthi, and occasional necrotic leaves on potato
- PVY O: mottling and mosaic symptom on tobacco and mild to
severe mosaic and leaf drop on potato
- PVY C: induces triple streak symptoms on some potato cultivars
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

18. PVY symptoms ■ PVY O ■ PVY N and PVY NTN
- causes a green and yellow mosaic
pattern in leaves, and/or as roughness
in the leaves in most hosts
- yield reduction, but no tuber quality loss
■ PVY N and PVY NTN
- necrotic strains which both cause veinal
necrosis on tobacco
- causes superficial necrotic ringspots on
infected tubers in certain potato varieties
- necrosis is referred to as potato tuber
necrotic ringspot disesase (PTNRD)
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

19. PVY infectivity in pepper (previous work)
■ Pepper, tobacco and potato isolates have their own classification based on biological criteria
■ PVY infectivity in pepper
- Strains were classified into 3 pathotypes based on their ability to overcome certain resistance genes
PVY - 0
PVY-1
PVY-1-2
pvr1 R
pvr11 S
pvr12
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.

20. Pepper infectivity for potato PVY strains
PVY N
PVY NO
PVY NTN
PVY O
Pvr1+ S R
pvr1
pvr11
pvr12
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.
- 5 Plants per genotype were screened.
- Pvr1+ and 3 different lines containing the pvr1 gene were screened.

21. Potato infectivity
■ Sandra or Chuck Brown sent 10 plants per cultivar which were
screened for resistance to the 4 PVY strains listed.
PVY N
PVY NO
PVY NTN
PVY O
Superior S
Russet Burbank
Dark Red
Norland
½ R
Umatilla
Katahdin
Tubers from these infected plants were harvested and are currently in cold storage to allow for symptom development. They will be screened for tuber necrosis in a few weeks.

22. pvr1 Transgenic Potatoes
Collaboration with Dr. Sandra Austin-Phillips - UW Madison
■ Constructs - Sent 1/05:
Pvr1+ - susceptible allele derived from C. annuum
pvr1 – resistant allele derived from C. chinense
pvr11 – resistant allele derived from C. annuum
pvr12 - resistant allele derived from C. annuum
■ Potato cultivars being transformed:
Katahdin
Russet Burbank
Superior
Umatilla (supplied by Chuck Brown)
Dark Red Norland

23. Transgenics potatoes (as of 12/05)
■ Independent transgenic events received:
■ Received 1-2 plants per event (Confirmed by Sandra via nptII PCR).
Pvr1
pvr1
pvr11
pvr12
Russet
Burbank 5 8 3
Katahdin 10

24. Summary and Future Plans
■ All potato varieties for transformation were found to be susceptible to all PVY N strains screened – PVY N, NO and NTN. Tubers of these infected plants are currently in cold storage to allow for PTNRD symptom development – they will be screened for tuber necrosis in a few weeks. ■ All pepper lines containing one of the 3 pvr1 resistance alleles: pvr1, pvr11 and pvr12 were found to be resistance to all 3 PVY N strains, while susceptible allele Pvr1+ was susceptible to both PVY N and PVY NTN. ■ All successful transformation events have been received up to this point They will be confirmed for transgene copy number as well as for transgene expression. ■ Cuttings will be made of all single copy transformants which will be subsequently screened for resistance to PVY O, N, NO and NTN.
Just to give you a big picture of my talk.
At first, I’m going to talk about a brief history of cop breeding in order to get you oriented.
An then I am going to talk about my work to date which is mostly on pepper.
After the research talk, I’ll be describing my future plans for education and research in Korean.