Identification of regions of cassava genome associated
with increased carotenoids content in roots
Ana Cruz Morillo, Alba Lucía Chávez, Jaime Marín, Martin Fregene, Luis Augusto Becerra López-Lavalle, Hernán Ceballos
International Center for Tropical Agriculture (CIAT). Cassava Breeding Project.
Mailing address 6713. Cali, Colombia. E-mail of contact person: l.a.becerra@cgiar.org
Introduction Table2. Correlation analysis between contrasting bulks (Yellow vs. White roots)
GM708 GM734 CM9816
Effects of vitamin A deficiency ranges from night blindness to Marker % Correlation Marker % Correlation Marker % Correlation
those of xerophthalmia and keratomalacia, leading to total blindness. SSRY-324 14 SSRY-200 10 SSRY-324 23
The principal vitamin A source is from animal origin, this has an SSRY-66 16 SSRY-9 10 SSRY-66 28
elevated cost for poor people. The pro-vitamin A carotenoids are SSRY-226 23 NS-267 10 SSRY-242 30
NS-267 26 SSRY-178 11 SSRY-172 33
cheaper source since they are found abundantly in plants. Cassava
SSRY-9 27 NS-119 14 SSRY-251 35
plays an important role as main source of low-cost energy for 800 SSRY-242 28 NS-53 17 SSRY-330 37
million of the world’s poorest people, but is not an important source SSRY-178 31 SSRY-21 18 NS-717 41
of other nutrients such iron, zinc, and vitamin A1. It has been SSRY-88 31 SSRY-66 27 SSRY-9 42
demonstrated that cassava germplasm with yellow roots may NS-717 32 SSRY-242 31 SSRY-195 42
SSRY-251 42 SSRY-313 35 NS-158 43
contribute significantly to resolve problem of Vitamin A deficiency in
SSRY-313 44 NS-717 41 SSRY-313 47
poor countries. We proposed to find molecular markers tightly linked SSRY-251 51
with high content of β-carotene to assist accelerating the breeding for
improved levels of β-carotene in cultivated cassava.
Population Phenotyping
Bulk
Materials and methods W W W W W
W
Plant material: thirty-two segregating families (756 ind.) derived from
reciprocal crosses between yellow, cream and white roots were grown and
evaluated for total carotene content. Y Y Y Y Y
Extraction and quantification of carotenoids: the methodology follows Y
standard protocols developed by HarvesPlus2.
Bulk segregant analysis (BSA): To find tightly linked markers with high
content of β-carotene, 800 microsatellites (SSRs) were screen between two
contrasting bulks (12 individuals each) for carotene content.
Figure 1. Bulk segregant analysis
Genetic Mapping: A genetic linkage map was constructed for cassava using
MapMaker/EXP version 3.0b3. Locus orders were
inferred with a LOD score of 3.0 using a G.1 G.2 G.3 G .4 G5
NS 980 rNS6 22 SSR Y25 0 3.9 rS SRY2 23 SS RY36
SS RY1 05
log-likelihood threshold of 2.0 and 2 0.3
4.4 NS G .1
717
SS RY9
24.1
19.9
SSRY 32
31.4
16.7
NS3 08
SSR Y20
1 0.9
2 3.0
NS 128 44.9
16.2
rN S158
1 5.8 rN S169
then using a log-likelihood threshold of 1.0.
rSSR Y95 32.0 NS 272 20.1
rS SRY2 51 10.2 rNS1 76 rN S124
1 0.3 5 .2 rNS8 0 NS1 89 2 5.5 15.0
NS 109 11.7 NS 149
SSRY 141 22.6 27.0
2 5.7 NS3 01 NS 376
21.5 SS RY57
32.7 2 2.0
rS SRY3 13 rNS9 09
19.9 NS3 84 SS RY1 77
2 5.2 12.2 SSR Y22 6
SSRY 183 G.10
15.1
Results
rS SRY6 6 SSRY 45 SSR Y39
17.9 G.9 22.6
3 4.0 rSSR Y241 rSSR Y256
26.9 G.8 r NS66 7 24.4
12.0
S SRY 324 rSSR Y55
4.0 SS RY92
rSS RY60 rNS1 78 rS SRY 7 1.2 S SRY 195 17.8
24.7 rSSR Y317
rS SRY 149 33.1
23.0
S SRY1 r SSRY 331
25.4
N S656
Table 1 show 5 of the 32 families selected
G.6 G7 G.1 1
23.7
r SSRY 64 SSR Y16 N S92 SS RY1 2
23.7
30.3 G .1 2 rN S945
28 .7 23.2
for the bulk segregant analysis (BSA). This lines 14.7 r SSRY 62 N S193
S SRY 47 G.1 3 NS1 36
rNS 658 13.9 23.2
29.8 NS3 41 rS SRY 31
21 .9 SS RY1 82 20.1
S SRY 205 2.4 NS2 71
were selected based on two criteria [high 26.7 3 .3 rNS 905
rNS 847 NS 170
8 .0 1.1
S SRY 172 rSS RY70 NS 207 G.15
G .1 4 SS RY18 1
carotenoid content and size of progeny (>80)]. The 32.7
G .1 9
SS RY41
14 .4
G.18
S SRY6 3
G.17
SSR Y278
32.3
rSSR Y21
rNS 1003
9.0
27 .9
rN S347
rSS RY2 10
SS RY27 2 rS SRY 178 16.5
bulks were selected as shown in Figure 1. The
23.6
NS1 94 rSSR Y81
G 20 G.21 G.22
results of the BSA analysis showed that SSR
G .1 6
SS RY11 7 SSR Y123 SSR Y242
6.4 23 .5 23.9 SS RY11 4
SS RY11 9 SSR Y143 15.5
NS1 19 NS 306
15.5
markers NS-717, SSRY-313 and SSRY-251 had the G 231 G24
G.25
rN S964
SS RY14 4 SS RY72
Figure 3. QTL mapping for high carotene content
highest degree of association with high carotene 31.7 3 0.1 SSR Y268
SS RY94 26.5
rS SRY1 67 rNS7 8
content (Table 2). For the QTL analysis, 229
individuals of the segregating population COL1684 Figure 2. Cassava genetic linkage map.
x MTAI1 were genotyped using 140 SSR markers and
the resulting genetic map had 25 linkage groups Table 3. Composite interval analysis for three carotene characteristics.
with a LOD of 4.0. The average distance between
markers is 21.5 cM covering 1,500 cM (Figure 2).
The linear regression analysis of the phenotypic
and genotypic data using MAPMAKERQTL (Figure
3), showed that NS-717, SSRY-313 and SSRY-251
were associated with three major QTLs for high
carotene content (Table 3).
Table 1. Segregating families with high content of carotenoids.
Root Root N° Total carotenoids
Crosses Female Male
colour colour individuals (ug/g FW)
CM 9816 MCOL 2295 Yellow SM 980-4 Cream 95 4.83 Conclusions
GM 705 MBRA 1A Yellow MCOL 1734 Cream 105 5.13
3.14 The BSA and the QTL analysis allow the identification of
GM 708 MBRA 1A Yellow MMAL 66 Cream 80
three SSR (NS-717, SSRY-313 and SSRY-251 ) putative associated
GM 734 MTAI2 Cream CM 3750-5 White 85 3.62
with high carotene content. These three markers are all located on
GM 893 SM 805-15 White MPER 297 Cream 108 3.10 linkage group D of the cassava genetic map.
References
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