3. Low temperature germination
protocol
A total of 39 BC4F2 bulks involving indica/japonica
crosses was subjected to LTG screening at 10oC for
19 d in a controlled growth chamber under dark
conditions using the method of Bertin et al. (1996)
with minor modifications.with minor modifications.
The experiment was carried out in two replications
by placing 50 sterilized seeds in a Petri plate, which
was immediately transferred into the growth
chamber after wetting the seeds with distilled water
equilibrated at 10 oC.
The number of germinated seeds per bulk was
recorded on 19 d after seeding.
4. Summary results of BC populations screened for low temperature germination
Details Low temperature germination
IR64 TQ NPT Total
Total BC2F2 populations 14 15 10 39
Single plant selections per BC
population 0 - 21 0 - 18 0 - 31
Total selected BC2F3 lines 79 77 51 207
Selection intensity (%) 5.64 5.13 5.10 5.31
Number of indica donors 1 1 0 2Number of indica donors 1 1 0 2
Selected lines 0 3 0 3
Selection intensity (%) 0.00 3.0 0.00 1.5
Number of japonica donors 9 11 7 27
Selected lines 55 52 39 146
Selection intensity (%) 6.11 4.73 5.57 5.41
Number of intermediate donors 4 3 3 10
Selected lines 24 22 12 58
Selection intensity (%) 6.00 7.33 4.00 5.80
For individual BC populations of 100 plants, a difference of 2.5% between two populations in selection intensity
(survival rate) is statistically significant at P < 0.05 when the selection intensity is < 0.1.
5. Screening for seedling cold tolerance
Twelve-day old seedlings were subjected to cold temperature for 18 days at the mean
daily temperature of 11.8 Co, including 3-day of low temperature at 8 Co between April
24-26 (LAAS, 2002).
6. Selection of 861 C418 plants with seedling cold tolerance
from 28 C418 BC2F2 populations 2002 (LAAS)
# of populations 28 2 26
BC2F2 CT donors
Non-CT
donors
Seedling Cold Tolerance (from NARES)
Range 1.4 – 19.3%
# of surviving plants
per population 10.3%
The mean population size was 310, ranging from 196 – 465, the recipient, C418 (japonica)
was killed by the stress.
10 – 16% 0 – 3.0%
0.314%
# of surviving plants
per BC population 10.3% 10.5%7.6%
7. Cold tolerance at the reproductive stageCold tolerance at the reproductive stage
8. Donor parent Subspecies Code
Population
size
No. of
selected
plants
SI 1
(%)
SF (%)
Mean ± SD2 Range
Bg90-2 Indica A 450 41 9.1 63.3±8.4D 50.3-86.5
X21 Indica B 450 29 6.4 64.3±9.9CD 50.6-87.1
X22 Indica C 450 28 6.2 65.6±10.1BCD 50.7-87.3
Screening results of 11 CY1 (recurrent parent) BC2F4 backcross populations for cold
tolerance at the booting stage in 2008 (1 SI = selection intensity, SF = spikelet fertility; 2
Different letters indicate statistical significance at P < 0.05, based on the Duncan
testing)
X22 Indica C 450 28 6.2 65.6±10.1BCD 50.7-87.3
Q5 Indica D 450 31 6.9 71.1±11.1ABC 50.9-91.4
Chhomrong Japonica E 450 24 5.3 75.6± 51.4-87.8
Doddi Indica F 450 25 5.6 71.0±10.0ABC 50.2-90.0
Fengaizhan Indica G 450 44 9.8 74.1± 52.2-98.5
Shennong265 Japonica H 450 21 4.7 69.2±11.4ABCD 50.7-93.8
Yuanjing7 Japonica I 450 41 9.1 71.1±9.8ABC 50.0-90.1
OM997 Indica J 450 21 4.7 72.1±9.1AB 54.9-89.6
Cs94 Indica K 450 19 4.2 64.2±8.6CD 51.4-86.0
Mean 450 29.5 6.5 69.2±4.2
CY1 Japonica 324 24.8±4.3E 19.0-30.0
9. Screening of the BC2F4 bulk populations for cold tolerance (CT) at the booting
stage:
Seeds of the BC2F4 bulk populations were sown in the seedling nursery on April 15,
2008, and 450 40-day old seedlings of each BC2F4 bulk population were transplanted
into a 45-row plot with 10 plants in each row and a spacing was 25×15cm in the
sheltered water pond of JAAS on May 25.
Two rows of CY1 were also transplanted on both sides of each plot as the checks.
•The water-pond were irrigated with water of normal temperature (25~28℃)
• When CY1 entered the stage of panicle initiation, the LT treatment was initiated
℃℃℃℃
℃℃℃℃
℃
• When CY1 entered the stage of panicle initiation, the LT treatment was initiated
by irrigation of flowing cold water (19±±±±0.5℃℃℃℃), which was adjusted in a nearby
water pool by mixing cold underground water (9℃℃℃℃) with the river water.
•The depth of the cold water in the pond was 20cm and the treatment was
maintained for ~30 days until panicles of almost all plants exerted completely.
•Then, irrigation with normal temperature water was resumed until the
maturity. At the maturity, all plants except those with >3 days earlier heading or
those with >3 days delayed heading than CY1 were harvested for measuring the
spikelet fertility.
•Under this LT treatment, CY1 had a spikelet fertility of 24.8±±±±4.3%, then, any
plant with spikelet fertility >50% were selected.
10. Donor parent
No. of
selected
lines
Cold stress at the seedling
stage Cold stress at the reproductive stage
Survival rate of seedlings
(%) SNP FGN SF (%)
Mean Range Mean Range Mean Range Mean Range
BG90-2 41 19.5 10.0~62.5 114.5 76.3~164.2 55.0 22.1~95.7 48.0 18.5~71.3
X21 29 19.4 11.1~40.3 99.7 73.3~125.0 50.5 29.5~90.3 50.3 30.9~74.9
X22 28 19.6 12.5~40.7 113.2 70.3~139.0 58.5 4.5~94.9 51.4 5.6~79.4
Q5 31 18.3 15.0~33.8 114.6 68.3~185.4 56.5 15.3~113.6 49.2 14.6~83.6
Evaluation of 324 BC2F6 introgression lines and their recurrent parent, CY1 for cold
tolerance of at the seedling and booting stages in 2009 (SNP = spikelet number per panicle,
FGN = filled grain number per panicle, SF = spikelet fertility)
Q5 31 18.3 15.0~33.8 114.6 68.3~185.4 56.5 15.3~113.6 49.2 14.6~83.6
Chhomromg 24 17.0 15.0~32.5 109.8 77.6~175.2 74.0 47.0~104.8 66.9 45.6~83.4
Doddi 25 17.3 12.5~25.0 112.9 89.4~182.6 73.9 51.5~108.5 65.4 48.7~80.6
Fengaizhan 44 16.7 12.5~25.0 96.5 70.2~115.1 67.1 14.3~96.0 70.1 12.7~86.1
Shennong265 21 15.6 0.0~23.3 96.9 65.0~144.8 62.0 36.9~99.2 63.0 40.0~79.9
Yuanjing7 41 18.5 15.0~45.0 116.0 93.5~151.0 80.6 30.4~106.4 69.2 25.5~84.2
OM997 21 37.1 15.0~82.4 81.6 56.2~108.1 42.0 14.9~64.0 51.1 23.3~72.0
Cs94 19 22.3 15.0~42.5 113.3 82.0~137.9 69.5 31.8~99.4 60.0 34.2~75.5
CY1 14.8 10.8~20.0 106.6 98.6~114.6 36.7 32.7~40.8 35.1 33.7~36.5
LSD0.05 4.5 8.3 8.2 6.2
