PhD

Blast disease, caused by Magnaporthe oryzae is one of the most
devastating diseases in rice and is a great threat to food security worldwide.
During kharif season, the disease is prevalent throughout the rice growing areas in
India including the southern states of Kerala, Tamil Nadu, Karnataka and Andhra
Pradesh. Approximately 100 major blast resistance genes have been reported in
rice and hence exploitation of host plant resistance through gene pyramiding can
be employed effectively for the management of blast. Developing Near Isogenic
Lines (NILs) ie., lines carrying each of the major resistance genes in the
background of susceptible recurrent parent is a major step in the pyramiding work
to be carried out for developing multi race resistant varieties. So, the present study
entitled “Development of Near Isogenic Lines of rice variety „Uma‟ for blast
resistance genes through molecular marker assisted backcross breeding” was
undertaken at the College of Agriculture, Vellayani Thiruvananthapuram, to
develop Near Isogenic lines (NILs) of rice variety Uma for blast resistance genes
(Pi1, Pi2 and Pikh) using identified donors through marker assisted back cross
breeding.
DNA markers closely linked to the blast resistance genes viz.,
RM527 (Pi2 gene), RM224 (Pi1 gene) and RM206 (Pikh gene) were used for
validating marker polymorphism in the identified traditional donors of blast
resistance genes viz. PTB21 (Thekkan) with Pi2, PTB7 (Parambuvattan) with Pi1
and Pikh and susceptible recipient parent Uma (MO16). This validation confirmed
the absence of genes in the recipient parent, „Uma‟. These polymorphic gene
specific markers were used for foreground selection in F1, BC1F1 and BC2F1
plants.
Hybridisation was carried out between recipient parent Uma and the two
donor parents viz., PTB21 and PTB7 to transfer genes for resistance. F1 plants
with heterozygous loci for blast resistance genes specific markers were identified
through foreground selection and backcrossed with „Uma‟ the recurrent parent to
obtain the BC1F1 generation. The BC1F1 plants were screened with foreground
markers and those plants containing the respective resistance genes were
identified. Among the plants screened for a particular cross, 12 were found to
contain Pi2 gene, 17 had Pi1 gene and 21 plants contained Pikh gene. χ2 test was
done with the genotypic data of BC1F1 plants with single genes viz. Pi1, Pi2 and
Pikh. Segregation ratio in BC1F1 population from all the crosses except
Uma x PTB7 (Pi1) did not show significant deviation from the expected 1:1 ratio
suggesting the presence of segregation distortion in the segregants of the cross
Uma x PTB7 (Pi1).
For the background selection, three parental lines were genotyped with 30
microsatellite loci and the ones polymorphic for Uma and the donors were
identified. Twelve markers were found to produce discrete amplicons for
distinguishing Uma and PTB21 while 17 markers could distinguish Uma and PTB7.
Five of the polymorphic markers were used for background selection in BC1F1
plants with resistance genes of the three crosses. The BC1F1 plants with highest
percentage of recurrent parent genome recovery in the three populations were
identified. Highest recovery percentage of 70 could be obtained in backcross
population of all the three crosses.
Morphological data for seven quantitative and one qualitative character
(kernel colour) was recorded for the BC1F1 plants with the resistant genes.
Euclidean coefficient of dissimilarity was assessed in comparison with the
recurrent parent Uma. The Euclidean distance values ranged from 5.9 (A-21) to
21.3 (A-9) in the cross Uma x PTB21 (Pi2), 4.88 (B-8) to 17.8 (B-23) in Uma x
PTB7 (Pi1) and from 4.79 (C-34) to 13 (C-15) in case of Uma x PTB7 (Pikh). All
the BC1F1 plants with resistance genes were used to raise the BC2F1 generation.
The BC2F1 populations of the three crosses were subjected to foreground
selection and plants with resistance genes were identified. Background selection
was carried out in BC2F1 plants with resistance genes using 12 polymorphic
markers in the cross Uma x PTB 21 (Pi2) and 17 markers in plants identified in
the BC2F1 of Uma x PTB7 (Pi1 and Pikh). Percentage of recurrent parent genome
recovery ranged from 70.83 to 95.83 for the cross Uma x PTB21 (Pi2), 70.59 to
97.06 for Uma x PTB7 (Pi1) and from 61.76 to 94.12 for Uma x PTB7 (Pikh).
Euclidean coefficient of dissimilarity was assessed in comparison with the
recurrent parent Uma, based on seven quantitative traits (plant height, number of
tillers number of productive tillers, panicle length, grain L/B ratio, thousand grain
weight and days to maturity) in the BC2F1 plants with resistance genes. The lowest
values of Euclidean distance were recorded in the plants A-16-6 (2.24), B-18-2
(3.35) and C-17-3 (4.33) of the three backcross populations respectively. Based on
the Euclidean distance and percentage recovery of the recurrent parent genome,
five plants were identified from each of the three crosses as potential Near
Isogenic Lines (NILs) for the three blast resistance genes Pi2, Pi1 and Pikh. The
genotype A-5-1 with a recurrent parent genome recovery of 95.83% and
Euclidean distance of 5.55 with Uma was identified as the most potential NIL of
Uma for the gene Pi2. Similarly B-27-1 (Euclidean distance – 4.56, % genome
recovery - 97.06) and C-17-3 (Euclidean distance - 4.33, % genome recovery -
94.12) were identified as potential NILs for Pi1 and Pikh genes respectively.
Selfed seeds (BC2F2) were collected from the identified potential NILs.
The present research could identify NILs of Uma for three blast resistance
genes viz., Pi1, Pi2 and Pikh with more than 94% recurrent parent genome
recovery with two backcrossing through marker assisted selection. This confirms
the utility of marker assisted backcross breeding in recurrent parent genome
recovery. The NILs can be used in intermating programmes in future to develop
pyramided lines of Uma with all the three blast resistance genes (Pi1, Pi2 and
Pikh) to ensure high yielding ability and durable broad spectrum resistance to the
blast pathogen.