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HORTSCIENCE 49(5):679–687. 2014.
Characterization and Performance of16 New Inbred Lines of LettuceIvan Simko4, Ryan J. Hayes, Carolee T. Bull, and Beiquan MouU.S. Department of Agriculture, Agricultural Research Service, U.S.Agricultural Research Station, 1636 E. Alisal Street, Salinas, CA 93905
Yaguang LuoU.S. Department of Agriculture, Agricultural Research Service, FoodQuality Laboratory, Henry A. Wallace Beltsville Agricultural ResearchCenter, Building 002, 10300 Baltimore Avenue, Beltsville, MD 20705
Mark A. Trent1, Amy J. Atallah, Edward J. Ryder2,and Rebecca G. Sideman3
U.S. Department of Agriculture, Agricultural Research Service, U.S.Agricultural Research Station, 1636 E. Alisal Street, Salinas, CA 93905
Additional index words. breeding, disease resistance, genetics, Lactuca sativa, postharvestquality, production, vegetable market
Lettuce (Lactuca sativa L.) is the mostpopular, commercially produced, leafy veg-etable in the world (FAOSTAT, 2013). Let-tuce cultivars are divided into horticulturaltypes based on the shape and size of the head;the shape, size, and texture of the leaves; andstem length (e.g., Simko, 2013). Three prev-alent types of lettuce used in U.S. cultivationare crisphead (which includes the subtypesiceberg and Batavia), romaine, and leaf.Crisphead cultivars form a spherical head;the iceberg subtype has a large, dense head,whereas the Batavia subtype has a smallerand less dense head. Romaine cultivars forman elongated head that may or may not closeon the top. Leaf-type cultivars are highlyvariable in terms of leaf color, shape, size,texture, leaf margin, and blistering. Leaf-typelettuces generally have leaves that are shorterthan romaine and heads that do not close tocover younger leaves.
The USDA lettuce breeding program inSalinas, CA, was initiated in 1956 (Whitaker,1974). The program has released a largenumber of highly popular cultivars, includingAutumn Gold, Climax, Merit, Pacific, Sali-nas, Salinas 88, Sea Green, Tiber, Vanguard,Vanguard 75, Winterset, etc. (COMPOSITdb,2013; Ryder, 1979a, 1979b, 1981, 1986, 1991;Ryder and Robinson, 1991; Ryder andWaycott, 1998; Thompson and Ryder, 1961).Most of these cultivars belong to the icebergtype, the major type of lettuce produced inthe United States through 1980s (for thesimplicity of description, the iceberg subtypewill be called ‘‘type’’ in the remaining part ofthe article). The USDA breeding priorities,however, shifted in the 1990s as a result of theincreased popularity of romaine and leaflettuces and also as a result of a larger in-volvement of private seed companies in let-tuce breeding. The USDA breeding program isfocused mostly on development of improvediceberg, romaine, and leaf lettuce inbred linesthrough introgression of desirable traits fromwild species, heirloom material, and un-adapted germplasm. The new lines are thenreleased into the public domain and used byprivate or public breeding programs directlyfor seed increase and sales or for developmentof new cultivars through additional rounds ofselection or mating.
We describe here the development andperformance of new iceberg (six), romaine(four), and leaf (six) breeding lines. Theseinbred lines were evaluated for performanceand resistance in field, greenhouse, and labo-ratory experiments. Detailed descriptions ofeach line provided here will allow seed com-panies to assess the suitability of the materialfor commercial production or for further de-velopment in their breeding programs.
Material and Methods
Field experiments. Eight field experi-ments were conducted in Salinas, CA, in
2012 and 2013 using a randomized com-plete block design with three replications(Table 1). Seeds were seeded in two rows35 cm apart on 102-cm wide beds (center-to-center), at least 7 m in length per genotype,and thinned to the final spacing of �30 cmbetween plants within a seedline, resulting in30+ plants per replicate. Crop cultivation wasdone using standard cultural practices for thearea except that fungicide treatment was notapplied to allow for assessment of diseaseresistance. Experiments included the 16 newinbred lines, their parents (or more distantancestors), and control cultivars. Because oflimiting factors such as the availability offield space, amount of seeds, disease prog-ress, or logistics, not all accessions wereevaluated for all traits in all experiments.The following accessions of the three horti-cultural types were evaluated: iceberg breed-ing lines RH08-0111, SM13-I1, SM13-I2,SM13-I3, SM13-I4, SM13-I5, parents [Calmar,Glacier, Iceberg (Batavia subtype), La Brillante(Batavia subtype), Salinas, Salinas 88] andcontrols (Silverado, Tiber); romaine breed-ing lines RH08-0464, SM13-R1, SM13-R2,SM13-R3, parents [Balady Banha (stem type),Darkland, Parris Island Cos, PI 491224] andcontrols (Green Towers, Hearts Delight,Lobjoits, Triple Threat); and leaf breedinglines SM13-L1, SM13-L2, SM13-L3,SM13-L4, SM13-L5, SM13-L6, parents(Grand Rapids, Lolla Rossa) and controls(Big Red, Big Star, Red Fox, Two Star).At least 12 of the evaluated parents andcontrols are current cultivars in production,including cvs. Big Red, Big Star, Darkland,Green Towers, Hearts Delight, Lolla Rossa,Parris Island Cos, Red Fox, Salinas (mar-keted under other names), Silverado, Tiber,and Two Star.
Horticultural traits, yield, and tipburn.Horticultural traits, yield, and tipburn wereevaluated at harvest maturity in the 12.5SPand 13.5SP experiments (Table 1). Ten plantsper plot that were representative of theaccession were harvested and weighed to-gether. An average head weight was deter-mined by dividing the total weight by 10.These 10 heads were cut in half vertically andthe number of heads with tipburn symptomswas recorded (as percentage). Five of theheads were used to measure core length, headheight (iceberg and romaine), head diameterat the top and bottom (romaine), and headdiameter at the midsection (iceberg). Themidsection head diameter was calculatedfrom two measurements of radii that wereperpendicular to each other. An experiencedevaluator visually estimated the number ofharvestable heads per plot. The heads wereconsidered harvestable if their size and shapemet the commercial standard for iceberg,romaine, or leaf lettuce. The percent ofharvestable heads was calculated from thetotal number of plants per plot. The sameevaluator assessed leaf ruffling and marginundulation and savoy, firmness, and closureof iceberg heads. Leaf ruffling and undulationand head savoy and closure were evaluatedon a scale from 1 to 4, where higher values
Received for publication 14 Feb. 2014. Acceptedfor publication 18 Mar. 2014.This research was supported by the CaliforniaLeafy Greens Research Program and the CaliforniaDepartment of Food and Agriculture SpecialtyCrop Block Grant Program SCB 10008.We thank Syngenta Seeds for seed increase ofadvanced breeding lines, Taylor Farms for trans-porting lettuce, and J. McCreight for reviewing themanuscript.Mention of trade names or commercial products inthis publication is solely for the purpose of pro-viding specific information and does not implyrecommendation or endorsement by the U.S. De-partment of Agriculture.1Current address: Pacific International Marketing,740 Airport Boulevard, Salinas, CA 93901,2Current address: 17739 Riverbend Road, SalinasCA 93908.3Current address: Department of Biological Sci-ences, University of New Hampshire, Durham, NH03824.4To whom reprint requests should be addressed;e-mail [email protected].
HORTSCIENCE VOL. 49(5) MAY 2014 679
indicated more pronounced characteristics.Head firmness was evaluated on a scale from1 to 5 (1 = no heading, 2 = closed, puffyheads, 3 = heads that slightly yield to handpressure, 4 = firm heads that do not yield tohand pressure, 5 = very firm, splitting heads).
Resistance to downy mildew. Six fieldexperiments (12.5SP, 12.6FA, 12.6SP,12.8FB, 13.5SP, and 13.8FB; Table 1) be-came naturally infected with Bremia lactu-cae. The resulting downy mildew severitywas evaluated at harvest maturity using a0 to 5 rating scale, where 0 = no lesions, 1 =sporadic lesions with less than one lesion perplant on average, 2 = up to two lesions perplant, 3 = three to 10 lesions per plant, 4 =more than 10 lesions per plant, and 5 = severeplant infection and merging of lesions(Simko et al., 2013). Field experiments didnot include accessions with resistance basedon functional R-gene(s), because our breedingprogram focuses on development of breedinglines with quantitative resistance to the disease(Simko, 2013; Simko et al., 2013).
Resistance to leafminer. Damage fromleafminers (Liriomyza sp.) resulting fromnatural infestations was evaluated in experi-ments 12.6FA, 12.6SP, and 12.8FB (Table 1).The total number of leafminer mines wascounted on two randomly selected plantsfrom each plot at the baby leaf stage (�30 dafter planting). Leafminer stings were evalu-ated at the baby leaf stage and at harvestmaturity. Stings within the 20-cm2 area of theleaf with the highest sting density werecounted for each plant per plot (Mou andLiu, 2003) and averaged.
Resistance to lettuce drop. Spring(12.5FB) and fall field experiments (12.8FBand 13.8FB) were conducted to evaluateresistance to lettuce drop caused by Scleroti-nia minor (Table 1). Plots were infested withsclerotia of S. minor in the spring of 2012 andagain in the spring of 2013 as described byHayes et al. (2010). For the 12.5FB experi-ment, sclerotia were applied immediatelybefore planting, whereas 12.8FB and 13.8FB
experiments assessed disease resistance in soilthat was infested with sclerotia produced fromthe previous crop. The percentage of plantsthat died from or exhibited lettuce drop symp-toms by market maturity of the iceberg culti-vars was recorded for each experiment.
Resistance to Verticillium wilt. Iceberg,romaine, and leaf lettuces were evaluatedfor resistance to Verticillium dahliae (race1) in a V. dahliae-infested field (12.5FC)according to methods of Hayes et al. (2007)(Table 1). At market maturity 10 plantsfrom each plot were uprooted. Disease in-cidence was assessed by cutting taprootslongitudinally and recording the number ofplants exhibiting any amount of discolor-ation of root vascular tissues typical ofVerticillium wilt.
Postharvest decay of fresh-cut lettuce.Lettuce grown in experiments 12.5SP, 12.6FA,12.6SP, and 13.5SP was used in evaluation ofdecay after processing (Table 1). Three heads(or more if heads were small) from each plotwere harvested from each of three replica-tions and placed in a 4 �C cold storage roomfor 1 d before processing. All the lettuceheads within an accession were bulked andprocessed into cut lettuce using the method ofHayes and Liu (2008). Nine bags per acces-sion were produced, triple-flushed with N2
gas, sealed, and stored at 4 �C. Each of the22.8 · 30.5-cm transparent bags contained340 g of tissue cut into 2.5-cm2 pieces. Decaywas visually evaluated in weekly intervalson a 0 to 10 scale that corresponds to theestimated percentage of decayed tissue di-vided by 10 (Simko et al., 2012). Theexperiment was ended 4 weeks after process-ing and individual ratings were combinedinto an overall score by the area under thedisease (deterioration in this analysis) prog-ress stairs approach (AUDPS) (Simko et al.,2012; Simko and Piepho, 2012).
Postharvest decay of whole heads. Let-tuce heads were harvested from 12.5SPand 13.5SP experiments for evaluation ofwhole-head shelf life. Romaine and leaf types
were harvested at peak maturity of ‘GreenTowers’, whereas iceberg types were har-vested at peak maturity of ‘Salinas’. Equalnumbers of heads from each replicate werepooled into a single carton, resulting in onecarton per breeding line or cultivar. Cartonswere cooled to 5 �C and shipped to Marylandthrough a refrigerated shipping truck pro-vided by Taylor Farms (Salinas, CA). Onarrival, the products were immediately trans-ferred to the USDA-ARS Food Quality Lab-oratory in Beltsville, MD, and stored at 5 �C.Lettuce head quality was evaluated for eachaccession at Day 0 (arrival quality), Day 7,Day 14, and for romaine and leaf accessionsalso at Day 17. After this period all thematerial was considered unacceptable forsale. Product quality was evaluated by atleast three trained evaluators. Overall qualitywas assessed following a modified procedurefrom Loaiza and Cantwell (1997) using a 9-point hedonic scale where 9 = like extremely;7 = like moderately; 5 = neither like nordislike; 3 = dislike moderately; and 1 =dislike extremely (Meilgaard et al., 1991).Scores from weekly evaluations were com-bined into an overall rating using the AUDPSapproach (Simko and Piepho, 2012). TheAUDPS scores were then converted into acomplementary scale (cAUDPS) by subtract-ing actual AUDPS scores from the maximumpossible AUDPSMax scores (rating of 9 at alldays) for the test: cAUDPS = AUDPSMax –AUDPS. This conversion was performed tokeep the scale consistent with scales for othertraits (e.g., resistance to diseases, tipburn, de-cays of fresh-cut lettuce, decay index) wherehigher values are less desirable.
Decay index was evaluated on the finalday when the majority of lettuce heads hadsome amount of decay. Lettuce heads foreach accession were cut in half and scores(0 to 4) were assigned based on the decayedareas: 0 = no decay; 1 = decayed area lessthan 25% of lettuce head; 2 = decayed arearanging from 25% to 50%; 3 = decayedarea ranging from 50% to 75%; 4 = decayed
Table 1. List of experiments, evaluated traits, and measurement units.
Expt. Location Planting date Evaluations and units
12.5FB Salinas—Field B 2 May 2012 Lettuce drop (%)12.5FC Salinas—Field C 5 May 2012 Verticillium wilt (%)12.5SP Salinas—Spence 8 May 2012 Head weight (g), head height (cm), head midsection diameter for iceberg (cm), head top diameter for
romaine (cm), head bottom diameter for romaine (cm), core length (cm), head closure (1 to 4 scale),tipburn (%), downy mildew (0 to 5 scale), salad decay (AUDPS), whole head decay (cAUDPS)
12.6FA Salinas—Field A 26 June 2012 Downy mildew (0 to 5 scale), leafminer stings at 30 d after planting (count), leafminer mines at 30 d afterplanting (count), leafminer stings at harvest maturity (count), salad decay (AUDPS)
12.6SP Salinas—Spence 26 June 2012 Downy mildew (0 to 5 scale), leafminer stings at 30 d after planting (count), leafminer mines at 30 d afterplanting (count), leafminer stings at harvest maturity (count), salad decay (AUDPS)
12.8FB Salinas—Field B 15 Aug. 2012 Downy mildew (0 to 5 scale), lettuce drop (%), leafminer stings at 30 d after planting (count), leafminermines at 30 d after planting (count), leafminer stings at harvest maturity (count)
13.5SP Salinas—Spence 13 May 2013 Head weight (g), head height (cm), head midsection diameter for iceberg (cm), head top diameterfor romaine (cm), core length (cm), head firmness for iceberg (1 to 5 scale), head savoy (1 to 4 scale),number of harvestable heads (%), tipburn (%), downy mildew (0 to 5 scale), salad decay (AUDPS),whole head decay (cAUDPS), decay index (0 to 1 range)
13.8FB Salinas—Field B 7 Aug. 2013 Downy mildew (0 to 5 scale), lettuce drop (%)GH Greenhouse 2013 Bacterial leaf spot (0 to 3 scale, transformed to 0 to 1 scale through integrated rating where 0 was given
to the most resistant accession from all horticultural types and the value of 1 was given to the mostsusceptible one)
Laboratory Laboratory 2013 Dieback (detection of resistant [R1, R2] or susceptible [S1] haplotypes with molecular markers)
AUDPS = area under the disease progress stairs approach; cAUDPS = complementary area under the disease (deterioration in this analysis) progressstairs approach.
680 HORTSCIENCE VOL. 49(5) MAY 2014
Tab
le2
.P
erfo
rman
ceo
fsi
xn
ewic
eber
gb
reed
ing
lin
esan
dei
gh
tco
mm
erci
alst
anda
rds
and
par
enta
lac
cess
ion
sas
mea
sure
db
y1
9q
ual
ity
and
dis
ease
reac
tio
ntr
aits
.
Tra
it(u
nit
)E
xp
t.M
ean
x
Com
mer
cial
stan
dar
ds
and
par
enta
lac
cess
ions
New
iceb
erg
lines
z
Cal
mar
Gla
cier
Iceb
erg
La
Bri
llan
teS
alin
asS
alin
as8
8S
ilv
erad
oT
iber
RH
08
-01
11
SM
13
-I1
SM
13
-I2
SM
13
-I3
SM
13
-I4
SM
13
-I5
Hea
dw
eig
ht(g
)1
2.5
SP
12
89
13
74
14
68
Hw
11
95
10
49L
13
05
14
05
—1
34
31
15
31
22
31
33
81
23
61
33
21
33
11
3.5
SP
90
61
06
61
05
07
59
65
7L9
84
93
69
53
88
46
15
L9
03
91
09
87
10
02
98
4H
ead
hei
ght
(cm
)1
2.5
SP
13
.71
4.0
13
.71
5.6
H1
7.4
H1
2.8
13
.8—
12
.81
2.7
13
.61
2.9
12
.91
3.4
12
.61
3.5
SP
16
.21
5.2
15
.42
2.7
H2
1.2
H1
5.6
15
.71
6.5
15
.01
4.6
14
.91
4.8
15
.01
5.9
14
.1H
ead
dia
met
er(c
m)
12
.5S
P1
5.9
15
.91
8.3
H1
7.0
18
.7H
15
.41
5.5
—1
4.2
14
.41
4.7
16
.31
5.0
15
.71
5.8
13
.5S
P1
6.6
17
.01
6.7
17
.81
6.9
16
.11
6.9
16
.91
6.5
16
.21
6.0
15
.21
6.5
17
.21
6.5
Co
rele
ngt
h(c
m)
12
.5S
P3
.32
.64
.1H
4.6
H3
.23
.23
.4—
2.9
3.1
3.0
3.6
3.0
3.2
3.5
13
.5S
P4
.74
.05
.46
.7H
5.4
4.3
4.5
4.2
3.6
3.6
4.6
5.0
4.8
5.3
5.0
Hea
dcl
osu
re(1
to4
)1
2.5
SP
3.9
4.0
4.0
3.5
L3
.7L
4.0
4.0
—4
.04
.04
.04
.04
.04
.04
.0H
ead
firm
nes
s(1
to5
)1
3.5
SP
3.3
4.0
3.3
1.3
L1
.0L
4.0
3.7
4.0
3.0
2.7
L3
.34
.03
.74
.04
.0H
ead
sav
oy
(1to
4)
13
.5S
P2
.62
.72
.73
.7H
3.0
2.0
2.0
2.0
2.0
2.0
3.0
3.3
H2
.73
.02
.7H
arv
esta
ble
hea
ds
(%)
13
.5S
P5
65
74
9—
—6
26
03
84
71
2L
71
75
68
80
H5
6T
ipb
urn
(%)
12
.5S
P3
32
37
3H
97
H9
7H
10
3—
10
33
33
17
17
20
0L
13
.5S
P5
73
37
78
71
00H
17
27
7L
7L
67
87
77
80
83
50
Do
wn
ym
ild
ew(0
to5
)1
2.5
SP
2.3
3.2
H3
.8H
0.7
L0
.6L
3.2
3.4
H—
3.4
H0
L2
.62
.42
.52
.42
.01
2.6
FA
2.8
2.8
4.0
H1
.91
.83
.23
.7H
——
1.5
3.5
2.7
3.0
2.8
3.0
12
.6S
P2
.1—
3.5
H1
.4L
0.6
L3
.3H
——
—1
.4L
2.5
H—
——
1.9
12
.8F
B2
.73
.34
.0H
1.8
1.5
L3
.53
.0—
3.8
H1
.82
.52
.32
.82
.52
.51
3.5
SP
2.9
3.3
4.0
H1
.6L
1.8
L3
.33
.7H
3.8
H3
.31
.3L
2.9
2.8
2.8
2.8
2.8
13
.8F
B3
.63
.84
.5H
2.3
L1
.9L
4.5
H4
.24
.4H
4.3
H2
.2L
4.0
3.5
3.7
3.8
3.3
Let
tuce
dro
p(%
)1
2.5
FB
58
37
57
52
54
55
46
—8
07
35
15
35
66
07
61
2.8
FB
24
7L
33
15
33
36
28
—3
03
61
31
41
32
13
01
3.8
FB
29
26
35
24
34
27
31
28
40
29
29
22
18
22
34
Lea
fmin
erst
ings
atD
ay3
0(c
ou
nt)
12
.6F
A1
4.1
14
.01
7.9
5.7
L2
.7L
18
.21
7.5
——
13
.41
8.4
15
.91
5.7
13
.41
6.0
12
.6S
P1
3.3
——
5.0
L—
——
——
10
.31
5.7
——
—2
2.3
H
12
.8F
B1
4.1
——
9.4
L1
1.9
——
——
15
.11
6.5
——
—1
7.4
Lea
fmin
erm
ines
atD
ay3
0(c
ou
nt)
12
.6F
A2
.72
.53
.3H
3.0
2.5
2.8
2.5
——
2.4
2.8
2.8
3.0
2.0
3.0
12
.6S
P2
.1—
—2
.0—
——
——
1.9
2.3
——
—2
.31
2.8
FB
2.4
——
2.2
2.3
——
——
2.3
2.3
——
—2
.7L
eafm
iner
stin
gs
ath
arv
est
(co
un
t)1
2.6
FA
11
.81
3.7
12
.55
.4L
3.2
L1
5.3
15
.8H
——
9.1
13
.51
3.2
14
.11
4.6
11
.0
12
.6S
P1
3.2
——
8.7
L—
——
——
14
.11
3.5
——
—1
6.6
H
12
.8F
B1
0.6
——
5.6
L6
.5L
——
——
13
.8H
12
.9H
——
—1
4.2
H
Bac
teri
alle
afsp
ot
(0to
1y)
GH
0.4
90
.55
0.5
60
.50
0.0
3L
0.6
00
.61
0.5
80
.57
0L
0.6
00
.54
0.5
80
.56
0.5
2S
alad
dec
ay(A
UD
PS
)1
2.5
SP
22
53
L3
2H
18
3H1
L2
L—
81
61
11
02
L5
51
2.6
FA
45
37
33
34
21
8H1
91
7—
—9
1H
18
32
13
L9
L1
81
2.6
SP
61
——
36
18
4H—
——
—5
01
7L
——
—1
9L
13
.5S
P3
32
24
L8
2H
24
0H4
L7
5L
19
61
11
42
16
14
Wh
ole
hea
dd
ecay
(cA
UD
PS
)1
2.5
SP
58
72
H6
08
4H
81
H4
6L
——
61
60
46
L3
7L
61
30
L6
1
13
.5S
P7
37
47
69
19
6H
74
62
71
47
61
63
96
H7
67
07
1D
ecay
inde
x(0
to1
)1
3.5
SP
0.4
70
.39
0.4
00
.97
H0
.90
H0
.29
0.3
10
.36
0.2
50
.37
0.4
00
.56
H0
.58
H0
.44
0.3
7D
ieb
ack
(hap
loty
pe)
Lab
ora
tory
—R
1R
1R
1R
1R
1R
1—
R1
R1
R1
R1
R1
R1
R1
zP
edig
rees
of
iceb
erg
bre
edin
gli
nes
incl
ud
ecv
s.C
alm
ar,
Gla
cier
,Ic
eber
g,
La
Bri
llan
te,
Sal
inas
,an
dS
alin
as8
8(d
etai
led
info
rmat
ion
isin
Tab
le6).
yO
rigin
alsc
ale
of
0to
3w
astr
ansf
orm
edto
0to
1th
rough
inte
gra
ted
rati
ng,
wher
e0
was
giv
ento
the
mos
tre
sist
ant
acce
ssio
n(f
rom
all
hort
icult
ural
typ
es)
and
the
val
ueo
f1
was
giv
ento
the
mo
stsu
scep
tib
leo
ne.
xM
ean
of
all
bre
edin
gli
nes
,co
ntr
ols,
and
par
ents
test
edin
the
exper
imen
t.wH
and
Lle
tter
sin
dic
ate
val
ues
sig
nifi
can
tly
(P<
0.0
5)h
igh
ero
rlo
wer
,re
spec
tiv
ely
,th
anth
eo
ver
all
mea
n.A
UD
PS
=ar
eaunder
the
dis
ease
pro
gre
ssst
airs
appro
ach;
cAU
DP
S=
com
ple
men
tary
area
under
the
dis
ease
(det
erio
rati
on
inth
isan
alys
is)
pro
gre
ssst
airs
app
roac
h.
HORTSCIENCE VOL. 49(5) MAY 2014 681
Tab
le3
.P
erfo
rman
ceo
ffo
ur
new
rom
ain
eb
reed
ing
lin
esan
dei
ght
com
mer
cial
stan
dar
ds
and
par
enta
lac
cess
ion
sas
mea
sure
db
y1
9q
ual
ity
and
dis
ease
reac
tio
ntr
aits
.
Tra
it(u
nit
)E
xp
t.M
ean
x
Com
mer
cial
stan
dard
san
dpar
enta
lac
cess
ions
New
rom
aine
lines
z
Bal
ady
Ban
haD
ark
lan
dG
reen
To
wer
sH
eart
sD
elig
ht
Lo
bjo
its
Par
ris
Isla
nd
Co
sP
I4
91
224
Tri
ple
Th
reat
RH
08
-04
64S
M1
3-R
1S
M1
3-R
2S
M1
3-R
3
Hea
dw
eig
ht(g
)1
2.5
SP
14
13
12
60
Lw
14
37
13
19
14
07
15
50H
13
78
13
38
—1
51
31
33
71
54
9H1
45
11
3.5
SP
10
12
88
61
02
21
00
21
12
01
11
39
02
87
21
01
31
17
29
61
97
21
10
7H
ead
hei
ght
(cm
)1
2.5
SP
29
.63
4.8
H2
7.7
28
.92
9.8
29
.02
8.7
29
.6—
31
.7H
29
.32
7.8
L2
8.5
13
.5S
P3
0.9
35
.9H
29
.92
8.4
L3
1.2
29
.73
0.2
28
.4L
32
.23
4.2
H3
1.5
29
.13
0.0
Hea
dto
pd
iam
eter
(cm
)1
2.5
SP
19
.51
6.1
L2
3.4
H1
8.3
16
.1L
21
.42
0.1
18
.4—
17
.52
0.4
22
.6H
19
.71
3.5
SP
29
.13
1.2
27
.63
0.9
30
.53
1.1
27
.02
9.6
29
.33
2.0
26
.92
4.8
28
.0H
ead
bo
tto
md
iam
eter
(cm
)1
2.5
SP
11
.49
.1L
11
.41
2.0
10
.61
3.7
H1
1.1
12
.3—
10
L1
0.3
12
.11
2.5
Co
rele
ngt
h(c
m)
12
.5S
P7
.21
0.6
H5
.0L
4.8
L5
.1L
9.9
H5
.2L
6.4
—9
.4H
9.0
H8
.9H
5.0
L
13
.5S
P8
.51
0.9
H7
.26
.9L
8.2
8.6
7.0
7.2
7.4
11
.6H
9.0
9.8
8.0
Hea
dcl
osu
re(1
to4
)1
2.5
SP
2.6
2.3
1.8
2.7
3.0
3.3
H2
.32
.5—
3.3
H2
.32
.52
.7H
ead
sav
oy
(1to
4)
13
.5S
P2
.83
.03
.03
.02
.33
.02
.72
.73
.02
.03
.03
.3H
2.0
Har
ves
tab
leh
ead
s(%
)1
3.5
SP
42
18
L6
2H
55
39
20
L6
8H
26
49
17
L5
05
04
5T
ipb
urn
(%)
12
.5S
P7
09
7H
53
50
77
93
67
67
—1
00
H5
07
34
31
3.5
SP
39
80
H1
31
05
35
73
32
75
08
3H
75
33
L
Do
wn
ym
ild
ew(0
to5
)1
2.5
SP
2.7
1.3
L2
.72
.53
.03
.7H
2.3
3.9
H3
.40
.8L
3.1
2.9
2.9
12
.6F
A3
.32
.83
.53
.43
.63
.33
.42
.93
.71
.93
.43
.83
.51
2.6
SP
2.4
——
2.5
3.1
H—
2.3
—2
.91
.0L
——
—1
2.8
FB
3.0
2.5
3.0
3.1
3.1
3.0
3.0
3.8
H3
.32
.3L
3.3
3.0
3.0
13
.5S
P2
.92
.22
.92
.83
.32
.82
.92
.93
.41
.43
.32
.93
.41
3.8
FB
3.8
2.7
L4
.5H
4.0
4.0
4.3
4.2
3.9
—2
.3L
4.3
4.2
3.7
Let
tuce
dro
p(%
)1
2.5
FB
58
54
55
73
64
57
56
37
—7
36
74
75
81
2.8
FB
36
20
43
56
H4
8—
38
24
40
38
43
19
27
13
.8F
B3
13
03
84
02
75
3H
41
21
—2
62
82
12
0L
eafm
iner
stin
gs
atD
ay3
0(c
ou
nt)
12
.6F
A1
1.7
8.7
13
.71
0.1
15
.28
.36
.9L
16
.5H
9.7
12
.41
2.7
8.6
18
.0H
12
.6S
P9
.1—
——
9.8
—9
.7—
5.6
L1
1.4
——
—1
2.8
FB
10
.8—
——
9.5
—1
0.9
—9
.81
3.0
——
—L
eafm
iner
min
esat
Day
30
(co
un
t)1
2.6
FA
2.4
2.3
2.3
2.3
2.3
2.8
2.3
2.8
2.5
2.0
2.3
1.8
L2
.5
12
.6S
P1
.9—
——
1.5
—1
.9—
2.5
1.6
——
—1
2.8
FB
2.2
——
—2
.2—
2.0
—2
.32
.2—
——
Lea
fmin
erst
ings
ath
arv
est
(co
un
t)1
2.6
FA
14
.01
2.9
14
.01
3.4
15
.31
3.9
10
.2L
18
.0H
12
.01
5.5
14
.31
2.3
16
.1H
12
.6S
P1
5.7
——
—1
6.3
—1
6.5
—1
2.4
17
.5—
——
12
.8F
B1
1.6
——
—1
0.3
—1
1.0
—1
1.7
13
.5—
——
Bac
teri
alle
afsp
ot
(0to
1y)
GH
0.6
21
.00H
0.6
30
.54
0.5
5—
0.5
50
.51
—0
.78H
0.5
70
.57
0.5
4S
alad
dec
ay(A
UD
PS
)1
2.5
SP
49
23
L1
8L
42
8L
38
11
L1
57H
—6
9H
13
6H
27
11
L
12
.6F
A8
51
23
19
L9
65
81
2L
51
18
7H1
90
H2
6L
18
5H
41
L2
6L
12
.6S
P7
9—
——
26
L—
36
L—
20
7H
45
——
—1
3.5
SP
68
34
25
L4
01
2L
20
L3
91
61H
15
2H
93
15
9H
68
23
L
Wh
ole
hea
dd
ecay
(cA
UD
PS
)1
2.5
SP
61
—3
9L
54
72
H—
65
40
L—
10
7H
60
72
H3
8.5
L
13
.5S
P6
19
5H
61
69
H5
24
9L
45
L5
75
47
3H
66
56
49
L
Dec
ayin
dex
(0to
1)
13
.5S
P0
.38
0.9
2H0
.28L
0.3
90
.24
L0
.25L
0.2
7L0
.43H
0.4
10
.51H
0.2
7L0
.29L
0.2
8L
Die
bac
k(h
aplo
typ
e)L
abo
rato
ry—
R2
S1
S1
S1
S1
S1
R2
R2
S1
R2
R2
R2
zP
edig
rees
of
rom
ain
eb
reed
ing
lin
esin
clu
de
cvs.
Bal
ady
Ban
ha,
Dar
kla
nd,
and
Par
ris
Isla
nd
Cos
and
acce
ssio
ns
PI
49
121
4an
dP
I4
91
224
(det
aile
din
form
atio
nis
inT
able
6).
yO
rigin
alsc
ale
of
0to
3w
astr
ansf
orm
edto
0to
1th
rough
inte
gra
ted
rati
ng,
wher
e0
was
giv
ento
the
mos
tre
sist
ant
acce
ssio
n(f
rom
all
hort
icult
ural
typ
es)
and
the
val
ueo
f1
was
giv
ento
the
mo
stsu
scep
tib
leo
ne.
xM
ean
of
all
bre
edin
gli
nes
,co
ntr
ols,
and
par
ents
test
edin
the
exper
imen
t.wH
and
Lle
tter
sin
dic
ate
val
ues
sig
nifi
can
tly
(P<
0.0
5)h
igh
ero
rlo
wer
,re
spec
tiv
ely
,th
anth
eo
ver
all
mea
n.A
UD
PS
=ar
eaunder
the
dis
ease
pro
gre
ssst
airs
appro
ach;
cAU
DP
S=
com
ple
men
tary
area
under
the
dis
ease
(det
erio
rati
on
inth
isan
alys
is)
pro
gre
ssst
airs
app
roac
h.
682 HORTSCIENCE VOL. 49(5) MAY 2014
Tab
le4
.P
erfo
rman
ceof
six
new
leaf
lett
uce
bre
edin
gli
nes
and
six
com
mer
cial
stan
dar
ds
and
par
enta
lac
cess
ions
asm
easu
red
by
17
qual
ity
and
dis
ease
reac
tion
trai
ts.
Tra
it(u
nit
)E
xp
erim
ent
Mea
nx
Com
mer
cial
stan
dard
san
dpar
enta
lac
cess
ions
New
leaf
lett
uce
lines
z
Big
Red
Big
Sta
rG
rand
Rap
ids
Lo
lla
Ro
ssa
Red
Fo
xT
wo
Sta
rS
M1
3-L
1S
M1
3-L
2S
M1
3-L
3S
M1
3-L
4S
M1
3-L
5S
M1
3-L
6
Hea
dw
eig
ht
(g)
12
.5S
P1
12
9—
—1
14
19
30L
w1
14
61
16
11
24
9H1
11
81
10
31
10
81
16
41
17
31
3.5
SP
73
87
63
76
06
76
33
4L8
04
80
78
72H
80
87
09
75
48
00
77
5H
ead
hei
ght
(cm
)1
2.5
SP
20
.2—
—1
7.9
L1
7.8
L2
4.2
H2
0.0
26
.2H
16
.81
7.3
L1
6.6
L2
2.0
23
.2H
Cor
ele
ng
th(c
m)
12
.5S
P6
.8—
—5
.29
.3H
6.5
4.0
L1
5.2
H6
.14
.3L
4.2
L6
.96
.31
3.5
SP
8.2
8.6
7.1
7.4
8.5
8.1
7.1
11
.3H
8.3
7.4
7.9
9.3
7.6
Hea
dcl
osu
re(1
to4
)1
2.5
SP
2.3
——
2.8
2.0
1.3
2.3
1.7
2.5
3.3
H3
.5H
1.8
1.8
Hea
dsa
vo
y(1
to4
)1
3.5
SP
3.3
3.0
2.3
L3
.32
.3L
3.3
2.7
4.0
4.0
4.0
4.0
3.7
3.3
Har
ves
table
hea
ds
(%)
13
.5S
P5
95
07
14
34
37
87
73
16
73
67
L7
27
8T
ipb
urn
(%)
12
.5S
P3
6—
—9
7H
0L
0L
33
10
27
70
H9
7H
10
15
13
.5S
P3
75
00
L7
01
01
03
71
03
L9
7H
10
0H2
04
0D
ow
ny
mil
dew
(0to
5)
12
.5S
P1
.13
.3H
—0
.80
.61
.42
.8H
0.9
0.2
L0
.2L
0.4
0.2
L1
.8H
12
.6F
A2
.02
.5—
2.2
1.7
—3
.7H
1.8
1.0
1.9
1.8
0.8
2.7
12
.6S
P1
.42
.4H
—0
.90
.5L
1.5
2.8
H1
.0—
1.0
—0
.8L
2.0
12
.8F
B2
.0—
—2
.01
.82
.03
.3H
1.3
L1
.51
.51
.51
.3L
2.8
H
13
.5S
P1
.62
.6H
2.1
1.3
0.9
2.4
H2
.6H
1.0
1.2
0.5
L1
.21
.41
.81
3.8
FB
2.4
3.3
H3
.4H
2.3
1.2
L2
.63
.6H
2.2
1.7
L1
.3L
2.0
1.6
L3
.3L
ettu
ced
rop
(%)
12
.5F
B5
4—
—3
95
46
96
45
03
15
05
77
15
31
2.8
FB
20
——
24
19
29
38
H4
L1
11
01
57
39
H
13
.8F
B2
62
24
71
72
53
73
01
31
71
91
91
84
2L
eafm
iner
stin
gs
atD
ay3
0(c
ou
nt)
12
.6F
A4
.38
.7H
—2
.53
.2—
6.6
H2
.42
.53
.24
.53
.35
.6
12
.6S
P4
.5—
—4
.4—
—8
.1H
2.2
—2
—4
.55
.71
2.8
FB
4.7
——
3.6
3.4
——
2.2
—4
.6—
—9
.5H
Lea
fmin
erm
ines
atD
ay3
0(c
ou
nt)
12
.6F
A2
.02
.8—
2.8
1.8
—2
.02
.32
.01
.82
.31
.80
.8L
12
.6S
P1
.7—
—2
.0—
—2
.9H
2.1
—1
.3—
1.4
0.6
L
12
.8F
B1
.6—
—1
.71
.8—
—2
.3—
1.2
——
1.0
Lea
fmin
erst
ing
sat
har
ves
t(c
oun
t)1
2.6
FA
4.7
10
.8H
—4
.92
.8—
5.9
2.5
3.0
2.6
2.3
3.9
7.9
12
.6S
P3
.9—
—3
.5—
—5
.5H
2.4
L—
2.5
L—
3.7
6.0
H
12
.8F
B3
.3—
—3
.32
.1—
—2
.0—
3.1
——
5.8
H
Bac
teri
alle
afsp
ot
(0to
1y)
GH
0.4
5—
—0
.56
0.5
40
.40
—0
.23
0.5
10
.39
0.5
50
.31
0.5
4S
alad
dec
ay(A
UD
PS
)1
2.5
SP
51
——
51
87
H7
9H
10
L4
44
47
3H
33
L5
73
5L
12
.6F
A8
71
56H
—4
9L
12
3—
41
L1
03
58
56
93
12
5H6
31
2.6
SP
69
12
5H—
46
71
78
97
H5
8—
46
—4
75
11
3.5
SP
94
90
43
L1
29H
65
11
46
4L
11
48
91
22
11
78
98
7W
ho
leh
ead
dec
ay(c
AU
DP
S)
12
.5S
P1
00
——
13
1H—
88
L1
19H
79
L1
02
11
0H
13
8H8
6L
51
L
13
.5S
P7
25
0L
47
L9
47
93
4L
68
70
71
10
5H
10
2H1
10H
37
L
Dec
ayin
dex
(0to
1)
13
.5S
P0
.48
0.4
90
.26L
0.7
1H0
.60
H0
.31L
0.5
80
.36L
0.3
5L
0.5
30
.53
0.6
9H0
.36
Die
bac
k(h
aplo
typ
e)L
abo
rato
ry—
——
R2
S1
—R
2S
1R
2R
2R
2R
2R
2zP
edig
rees
of
leaf
lett
uce
bre
edin
gli
nes
incl
ude
cvs.
Gra
ndR
apid
s,Ic
eber
g,
and
Loll
aR
oss
aan
dac
cess
ion
PI
49122
4(d
etai
led
info
rmat
ion
isin
Tab
le6
).yO
rigin
alsc
ale
of
0to
3w
astr
ansf
orm
edto
0to
1th
rough
inte
gra
ted
rati
ng,
wher
e0
was
giv
ento
the
most
resi
stan
tac
cess
ion
(fro
mal
lhort
icult
ura
lty
pes
)an
dth
ev
alu
eo
f1
was
giv
ento
the
mo
stsu
scep
tib
leo
ne.
xM
ean
of
all
bre
edin
gli
nes
,co
ntr
ols
,an
dp
aren
tste
sted
inth
eex
per
imen
t.wH
and
Lle
tter
sin
dic
ate
val
ues
sign
ifica
ntl
y(P
<0
.05)
hig
her
or
low
er,
resp
ecti
vel
y,
than
the
ov
eral
lm
ean.
AU
DP
S=
area
un
der
the
dis
ease
pro
gre
ssst
airs
app
roac
h;
cAU
DP
S=
com
ple
men
tary
area
un
der
the
dis
ease
(det
erio
rati
on
inth
isan
aly
sis)
pro
gre
ssst
airs
app
roac
h.
HORTSCIENCE VOL. 49(5) MAY 2014 683
area greater than 75%. Decay indices werecalculated according to the formula: DI = (0 ·N0 + 1 · N1 + 2 · N2 + 3 · N3 + 4 · N4) O(4 · NT), where N0 to N4 are the quantities oflettuce heads assigned each respective scoreand NT is the total number of lettuce headsexamined for the accession.
Resistance to bacterial leaf spot. Evalua-tions of seedlings for resistance to bacterialleaf spot caused by Xanthomonas campestrispv. vitians (Xcv) were conducted using agreenhouse assay that provides data that areclosely correlated with results in field exper-iments (Bull et al., 2007; Hayes et al., 2008,2014). Seeds were planted in plug trays(22 cm wide and 65 cm long) with 31 rowsof 11 cells that are 20 mm · 20 mm · 60 mmrepresenting a final plant density of �2380plants/m2. Plots consisted of a single row ofnine plants with the perimeter of each plugtray planted with the susceptible cv. VistaVerde. In the first of three experiments, threeXcv strains (BS339, BS340, and BS347) wereused as inoculum; however, after determin-ing that strain BS347 was more virulentthan the other strains (data not shown), onlyBS347 was used in the second and thirdexperiments. All of these strains were iso-lated from lettuce plants in California (Barakand Gilbertson, 2003; Bull and Koike, 2005).Xcv suspensions of �1 · 108 colony-formingunits/mL were prepared according to Bullet al. (2007). The suspensions of each isolatewere then mixed in equal proportions or thesingle isolate was used as prepared. Seedlingswere sprayed to runoff with the Xcv suspen-sion and incubated under constant leaf wet-ness for 7 d. Plants were rated for diseaseseverity using a 0 to 3 rating scale (0 = nodisease; 1 = few lesions less than 3 mm; 2 =lesions greater than 3 mm; 3 = coalescedlesions). Disease severity values were aver-aged across all plants within each treatmentreplicate. Data from multiple experimentswere combined into an overall integrated ratingusing rank-aggregation approach (Simko et al.,2012; Simko and Piepho, 2011). Integratedratings were then linearly adjusted to a 0 to 1scale, where the value of 0 was given tothe most resistant accession from all horti-cultural types and the value of 1 was given tothe most susceptible one.
Resistance to lettuce dieback. Lettucedieback is caused by two soilborne virusesfrom the family Tombusviridae: Tomatobushy stunt virus and Lettuce necrotic stuntvirus. Resistance to the disease is conferredby Tvr1, a single dominant gene (Grube et al.,2005; Simko et al., 2009). Resistance tolettuce dieback was evaluated with molecularmakers closely linked to the Tvr1 gene(Simko et al., 2009). Presence of the re-sistance alleles in lettuce accessions wastested by high-resolution DNA melting assayaccording to Simko (2013).
Statistical analyses. Data from each of thethree lettuce types were analyzed separatelywith the exception of the Verticillium wiltexperiment that combined all horticulturaltypes. Normality of data distribution wastested with Shapiro-Wilk W test implemented
in JMP 9.0.0 software (SAS Institute, Cary,NC). Traits with normally distributed datawere analyzed using analysis of means(ANOM). Traits with non-normally distrib-uted data or ratings performed on ordinalscales were analyzed with ANOM for ranks(JMP 9.0.0 software). The P value to identifyaccessions significantly different from theoverall mean was set at 0.05. Integratedratings from ranked data were calculated withWinsteps 3.65.0 (Winsteps.com, Beaverton,OR) according to Simko and Linacre (2010).
Results and Discussion
Five iceberg lines (SM13-I1 to SM13-I5)formed heads of acceptable quality withweight, height, diameter, core lengths, andfirmness similar to the overall mean andcontrol cultivars that represent industry stan-dard (Table 2). The percentage of harvestableheads for these five lines was also acceptablewith SM13-I4 having a significantly higherpercentage of harvestable heads per area thanthe overall mean. Tipburn incidence was gen-erally higher in the breeding lines comparedwith the tipburn-resistant control cvs. Silver-ado and Tiber; however, SM13-I5 showedno tipburn in 12.5SP. Resistance of SM13-I1through SM13-I5 to downy mildew wasusually better than in the established icebergcvs. Glacier, Salinas, Salinas 88, Silverado,and Tiber, the favorable alleles probablyoriginating from the highly resistant cv.Iceberg. Resistance to other diseases andpests (lettuce drop, bacterial leaf spot, andleafminer) was similar to the iceberg controlcultivars. Decay of fresh-cut lettuce andwhole head decay was similar to cv. Salinaswith the exception of SM13-I2 and SM13-I3,in which more rapid (but still acceptable)decay was observed on whole heads. RH08-0111 formed smaller and less firm heads thanestablished cultivars or other breeding lines;thus, the percentage of harvestable heads was
significantly smaller for this line. However,RH08-0111 had significantly higher resis-tance to downy mildew and bacterial leafspot compared with the overall averages.Alleles for the higher resistance to thesediseases in this breeding line likely originatefrom Batavia cv. La Brillante. All icebergbreeding lines possessed the R1-haplotypealleles indicating a complete resistance tolettuce dieback.
Three romaine breeding lines (SM13-R1to SM13-R3) had heads of average size andyield (Table 3). SM13-R3 had significantlylower tipburn incidence than the overall meanin one experiment (13.5SP). The remainingbreeding line (RH08-0464) formed elongatedheads with a long core, low percentage ofharvestable heads, and high tipburn inci-dence. Alleles for these traits were likelyinherited from cv. Balady Banha that is notused for commercial production in the UnitedStates. However, this breeding line had con-sistently less downy mildew than other testedromaine accessions (values were signifi-cantly different in four of six experiments).This breeding line had high susceptibility tobacterial leaf spot and more rapid decay ofwhole heads than was the mean of romaineaccessions. These traits were likely inheritedfrom cv. Balady Banha. RH08-0464 inheritedalleles for susceptibility to dieback from cv.Darkland. The other three breeding lines hadaverage resistance to downy mildew andbacterial leaf spot. SM13-R3 had a high num-ber of leafminer stings but good shelf life aswhole heads and fresh-cut lettuce. SM13-R1had poor shelf life when processed for saladbut acceptable shelf life of whole heads.SM13-R2 showed good resistance to leaf-miners and acceptable postharvest quality forwhole heads and fresh-cut lettuce. All threebreeding lines had the R2-haplotype allelesinherited from accessions PI 491224 or PI491214, indicating their complete resistanceto dieback.
Table 5. Reactions of new breeding lines, commercial standards and parental accessions to Verticilliumwilt race 1.
Accession Horticultural type Verticillium wilt incidence (%)
Commercial standards and parental accessionsBalady Banha Stem 27Darkland Romaine 57Hearts Delight Romaine 50La Brillante Batavia 0Ly
Lobjoits Romaine 47Lolla Rossa Leaf 0L
Parris Island Cos Romaine 54PI 491224 Romaine 60Red Fox Leaf 69Salinas Iceberg 83H
Tiber Iceberg 67New breeding linesz
RH08-0111 Iceberg 0L
RH08-0464 Romaine 87H
S13-R1 Romaine 57S13-R2 Romaine 73S13-R3 Romaine 43S13-L6 Leaf 0L
Mean 46zDetailed pedigree information is in Table 6.yH and L letters indicate values significantly (P < 0.05) higher or lower, respectively, than the overallmean.
684 HORTSCIENCE VOL. 49(5) MAY 2014
Tab
le6
.P
edig
ree,
des
crip
tio
n,
and
po
ten
tial
use
of
six
iceb
erg
,fo
ur
rom
ain
e,an
dsi
xle
afle
ttu
cen
ewb
reed
ing
lin
es.
Lin
eP
edig
ree
Des
crip
tion
Ad
van
tag
ezD
isad
van
tag
ezP
ote
nti
alu
se
RH
08
–0
111
F8:
Sal
inas
88
·L
aB
rill
ante
Gre
en,
slig
htl
yru
ffled
leav
esw
ith
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ew,
bac
teri
alle
afsp
ot,
and
Ver
tici
lliu
mw
ilt
race
1
Sm
all,
soft
hea
ds
that
do
no
tm
eet
the
mo
der
nic
eber
gst
anda
rdS
alad
ble
nd,
bre
edin
gp
rog
ram
s
SM
13
-I1
F6:
[(S
alin
as·
Iceb
erg)
·(I
ceb
erg
·C
alm
ar)]
·G
laci
erG
reen
,sl
igh
tly
ruffl
edle
aves
wit
hu
nd
ulat
ion
Imp
rov
edre
sist
ance
tod
ow
ny
mil
dew
Su
scep
tib
leto
tip
bu
rnW
ho
leh
ead
,sa
lad
ble
nd
SM
13
-I2
F6:
[(S
alin
as·
Iceb
erg)
·(I
ceb
erg
·C
alm
ar)]
·G
laci
erG
reen
,sl
igh
tly
ruffl
edle
aves
wit
hu
nd
ulat
ion
Imp
rov
edre
sist
ance
tod
ow
ny
mil
dew
Rel
ativ
ely
rap
idd
ecay
of
wh
ole
hea
ds
Wh
ole
hea
d,
sala
db
lend
SM
13
-I3
F6:
[(S
alin
as·
Iceb
erg)
·(I
ceb
erg
·C
alm
ar)]
·G
laci
erG
reen
,sl
igh
tly
ruffl
edle
aves
wit
hu
nd
ulat
ion
Imp
rov
edre
sist
ance
tod
ow
ny
mil
dew
Rel
ativ
ely
rap
idd
ecay
of
wh
ole
hea
ds
Wh
ole
hea
d,
sala
db
lend
SM
13
-I4
F6:
[(S
alin
as·
Iceb
erg)
·(I
ceb
erg
·C
alm
ar)]
·G
laci
erG
reen
,sl
igh
tly
ruffl
edle
aves
wit
hu
nd
ulat
ion
Hig
hper
centa
ge
of
har
ves
table
hea
ds,
imp
rov
edre
sist
ance
tod
ow
ny
mil
dew
Su
scep
tib
leto
tip
bu
rnW
ho
leh
ead
s,sa
lad
ble
nd
SM
13
-I5
F8:
(Ice
ber
g·
Sal
inas
)·
(Ice
ber
g·
Sal
inas
)G
reen
,sc
allo
ped
leav
esw
ith
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ewW
ho
leh
ead
s,sa
lad
ble
nd
RH
08
–0
464
F6:
Bal
ady
Ban
ha·
Dar
kla
nd
Dar
kg
reen
smo
oth
leav
esw
ith
slig
ht
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ewL
on
gco
re;
smal
lp
erce
nta
ge
of
har
ves
tab
leh
ead
s;su
scep
tible
toti
pburn
,bac
teri
alle
afsp
ot,
Ver
tici
lliu
mw
ilt
and
die
back
,ra
pid
dec
ayo
fw
ho
leh
ead
s
Bre
edin
gp
rog
ram
s
SM
13
-R1
F7:
PI
49
12
24
·P
arri
sIs
lan
dC
osG
reen
smo
oth
leav
esw
ith
un
dul
atio
nL
ow
tip
bu
rnin
cid
ence
,re
sist
ant
todie
bac
kR
apid
dec
ayaf
ter
pro
cess
ing
for
sala
dW
ho
leh
ead
SM
13
-R2
F8:
PI
49
12
24
·P
arri
sIs
lan
dC
osG
reen
smo
oth
leav
esw
ith
un
dul
atio
nS
lig
htly
mor
ere
sist
ant
tole
afm
iner
,sl
owd
ecay
so
fw
ho
leh
ead
s,re
sist
ant
todie
bac
k
Wh
ole
hea
d,
spri
ng
mix
,sa
lad
ble
nd
SM
13
-R3
F8:
Dar
kla
nd
·P
I4
91
21
4G
reen
smo
oth
leav
esw
ith
un
dul
atio
nL
ow
tip
bu
rnin
cid
ence
,sl
ow
dec
ayo
fw
ho
leh
ead
s,sl
ow
dec
ayaf
ter
pro
cess
ing
for
sala
d,
resi
stan
ceto
die
back
Su
scep
tib
leto
leaf
min
erW
ho
leh
ead
,sp
rin
gm
ix,
sala
db
lend
SM
13
-L1
F7:
Gra
ndR
apid
s·
Iceb
erg
Lig
ht
gre
enru
ffled
leav
esw
ith
un
dul
atio
nR
elat
ivel
yla
rge
hea
d,
imp
rove
dre
sist
ance
tod
ow
ny
mil
dew
,sl
owd
ecay
of
wh
ole
hea
ds
Lo
ng
core
(ear
lyb
olt
ing
),su
scep
tibl
eto
die
back
Wh
ole
hea
d,
spri
ng
mix
,sa
lad
ble
nd
SM
13
-L2
F7:
Gra
ndR
apid
s·
Iceb
erg
Lig
ht
gre
enru
ffled
leav
esw
ith
un
dul
atio
nL
ow
tip
bu
rnin
cid
ence
,im
pro
ved
resi
stan
ceto
do
wn
ym
ild
ew,
slow
dec
ayo
fw
ho
leh
ead
s,re
sist
ant
tod
ieba
ck
Wh
ole
hea
d,
spri
ng
mix
,sa
lad
ble
nd
SM
13
-L3
F7:
Gra
ndR
apid
s·
Iceb
erg
Lig
ht
gre
enru
ffled
leav
esw
ith
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ew,
resi
stan
tto
die
back
Su
scep
tib
leto
tip
bu
rn,
rela
tiv
ely
rap
idd
ecay
of
wh
ole
hea
ds
Sp
rin
gm
ix,
sala
db
lend
SM
13
-L4
F7:
Gra
ndR
apid
s·
Iceb
erg
Lig
ht
gre
enru
ffled
leav
esw
ith
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ew,
resi
stan
tto
die
back
Su
scep
tib
leto
tip
bu
rn,
low
per
cen
tag
eo
fhar
ves
tabl
ehea
ds,
rela
tivel
yra
pid
dec
ayo
fw
ho
leh
ead
s
Sp
rin
gm
ix,
sala
db
lend
SM
13
-L5
F7:
Gra
ndR
apid
s·
Iceb
erg
Lig
ht
gre
enru
ffled
leav
esw
ith
un
dul
atio
nIm
pro
ved
resi
stan
ceto
do
wn
ym
ild
ew,
resi
stan
tto
die
back
Rel
ativ
ely
rap
idd
ecay
of
wh
ole
hea
ds
Sp
rin
gm
ix,
sala
db
lend
SM
13
-L6
F7:
Lo
lla
Ro
ssa
·P
I4
91
224
Dar
kg
reen
/red
dish
smo
oth
leav
esw
ith
un
dul
atio
nS
low
dec
ayo
fw
ho
leh
ead
s,re
sist
ant
todie
back
and
Ver
tici
lliu
mw
ilt
race
1L
ess
resi
stan
tto
do
wn
ym
ilde
wth
ano
ther
fiv
en
ewb
reed
ing
lin
eso
fle
afle
ttu
ceW
ho
leh
ead
,sp
rin
gm
ix,
sala
db
lend
zA
dv
anta
ges
and
dis
adv
anta
ges
asco
mp
ared
wit
hco
mm
erci
alst
anda
rdcu
ltiv
ars
for
the
resp
ecti
ve
ho
rtic
ult
ura
lty
pe.
HORTSCIENCE VOL. 49(5) MAY 2014 685
Breeding lines of leaf lettuce producedheads of acceptable size (Table 4). SM13-L1had above average head weight accompaniedby longer cores, probably as a result of earliermaturity and the initiation of bolting. SM13-L2 had lower than average incidence oftipburn (in 13.5SP), whereas SM13-L4 andSM13-L5 had higher than average tipburnincidence. The low percentage of harvestableheads in SM13-L4 was caused by hightipburn incidence. Five leaf-type breedinglines (SM13-L1 to SM13-L5) had averageresistance to lettuce drop, bacterial leaf spot,and leafminers. Postharvest decay after pro-cessing for salad was in the acceptable range,although it was significantly higher than the
overall mean in two cases. Two of the lines(SM13-L1 and SM13L2) showed less rapiddecay of whole heads, whereas two lines(SM12-L3 and SM13-L4) had decay morerapid than the average. Results for SM13-L5were inconsistent. All five lines had averageto above average resistance to downy mil-dew. The resistance alleles in these linesoriginate from both the Batavia cv. Icebergand the leaf cv. Grand Rapids (Simko et al.,2013). The remaining breeding line (SM13-L6) was relatively more susceptible to downymildew and lettuce drop. Interestingly, thisline had often a relatively high number ofleafminer stings but very few leafminer mines.This may indicate presence of a limiting factor
that prevents leafminer laying eggs or de-velopment of larvae. SM13-L6 had acceptablepostharvest quality of whole heads and alsoafter processing for salad. All leaf-type breed-ing lines, with the exception of SM13-L1,had the R2-haplotype alleles inherited fromcv. Grand Rapids and PI 491224. Thesealleles are associated with the complete re-sistance to dieback.
Testing a selected subset of accessions in afield infected with V. dahliae race 1 (12.5FC)identified two resistant breeding lines (Table5). The iceberg breeding line RH08-0111inherited Vr1 resistance gene from cv. LaBrillante (Hayes et al., 2011), whereas theleaf-type breeding line SM13-L6 inherited
Fig. 1. Phenotypes of 16 released breeding lines from three horticultural types. The top four rows show plants growing in Expt. 12.5SP. The bottom row showsselected lines from the same experiment before shipping to Maryland for evaluations of whole head decay.
686 HORTSCIENCE VOL. 49(5) MAY 2014
resistance from cv. Lolla Rossa. It is unknownif the resistance in cv. Lolla Rossa is alsoconferred by Vr1. Breeding lines not tested forresistance to Verticillium wilt will likely havemoderate to high susceptibility to the disease,because all accessions in their pedigree aresusceptible to the disease (Hayes et al., 2007;Hayes, unpublished results and present data).
The 16 new breeding lines can be used fordifferent markets or breeding programs basedon their performance. The relative performanceof inbred lines (as compared with accessionsof the same type), their advantages, disad-vantages, and recommended use are sum-marized in Table 6, whereas phenotypes areshown in Figure 1. Five of six iceberg lines(SM13-I1, SM13-I2, SM13-I3, SM13-I4,SM13-I5) are suitable for salad blend andwhole head markets. Although SM13-I2and SM13-I3 exhibited more rapid decay ofwhole heads than the overall average, theirrates of decay are expected to be acceptablefor commercial production. RH08-0111 isnot acceptable for commercial productionof whole heads because of its small, softheads that do not meet the standard formodern iceberg cultivars. However, the linecan be used in iceberg breeding programs asa donor of alleles for resistance to downymildew, bacterial leaf spot, and Verticilliumwilt race 1 (Hayes et al., 2011). Two romainebreeding lines (SM13-R2 and SM13-R3) aresuitable for salad blend, spring mix, andwhole head production. Romaine breedingline SM13-R1 cannot be used for fresh-cutproducts, because it decays rapidly afterprocessing, but it is suitable for the wholehead market. An important trait of these threebreeding lines is their resistance to dieback,because most of the currently grown romainecultivars are susceptible to this disease(Simko et al., 2009). Romaine breeding lineRH08-0464 has improved field resistance todowny mildew inherited from cv. BaladyBanha. RH08-0464 can be used in breedingprograms but is not suitable for commercialproduction. All six leaf lettuce breeding linesare acceptable for commercial productionof salad blend and spring mix. SM13-L1,SM13-L2, and SM13-L6 could also be usedfor whole head production. SM13-L1 toSM13-L5 demonstrated very high field re-sistance to downy mildew with resistancealleles inherited from cvs. Iceberg and GrandRapids. Resistance to downy mildew in theselegacy cultivars appears to be durable as itwas reported over 50 years ago in cv. GrandRapids (Verhoeff, 1960) and over 90 yearsago in cv. Iceberg (Milbrath, 1923). How-ever, the lack of recent use of these cultivarsin a high-intensity production environmenthas reduced their exposure to sustained se-lection for virulent races of the pathogen.Five of the leaf lettuce breeding lines (SM13-L1 being the exception) have alleles for die-back resistance, a disease that may substantiallylimit lettuce production in California and Ari-zona (Simko et al., 2009). SM13-L1 is earlier
bolting than other breeding lines. SM13-L6has resistance to Verticillium wilt race 1.
On average �99.2% of loci that wereheterozygous in F1 are expected to be homo-zygous in F8. The value decreases to 98.4% inF7 and 96.9% in F6 generations. Therefore,there is a possibility that some of the releasedbreeding lines may be still genetically segre-gating for some of the traits. No segregationis expected, however, for resistance to die-back, because only lines homozygous at Tvr1were selected for seed production throughmarker-assisted selection.
Limited samples of seeds are available fordistribution to all interested parties for researchor commercial purposes. It is requested thatappropriate recognition be made if the breed-ing lines contribute to research or the devel-opment of new germplasm, breeding lines, orcultivars. Written requests should be sent tothe first or the second author.
Literature Cited
Barak, J.D. and R.L. Gilbertson. 2003. Geneticdiversity of Xanthomonas campestris pv.vitians, the causal agent of bacterial leafspotof lettuce. Phytopathology 93:596–603.
Bull, C.T., P.H. Goldman, R.J. Hayes, L.V. Madden,S.T. Koike, and E.J. Ryder. 2007. Geneticdiversity of lettuce for resistance to bacterialleaf spot caused by Xanthomonas campestrispv. vitians. Plant Health Prog. DOI: 10.1094/PHP-2007-0917-02-RS.
Bull, C.T. and S.T. Koike. 2005. Evaluating theefficacy of commercial products for managem-ant of bacterial leaf spot on lettuce. Plant HealthProg. DOI: 10.1094/PHP-2005-1121-01-RS.
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