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Exploitation of hybrid vigour is one of the most important applications of genetics in agriculture. It has highly contributed to food security, without affecting the environment. The availability of effective restorer lines that combines well with the CMS lines for various agro-morphological traits, determines the success of hybrid rice technology under temperate conditions like Kashmir Valley. To identify the best performing lines of rice in different cross combinations, four CMS lines and 34 genotypes were characterized for various agro-morphological and fertility restoration related traits. Out of 136 crosses, 43 (31.6%) were found to show the fertility restoration and 30 (22.06%) were found to be partial restorers. Sixty-three (46.32%) cross combinations were found to be partial maintainers and no cross combination was found to exhibit the maintainer behaviour. The genotypes RL-2, RL-3 and RL-8 were found to show restoration across all the CMS lines. Analysis of variance indicated the existence of significant differences among the genotypes, revealing that sufficient variability is present for different traits. The maximum positive value of st and ard heterosis for grain yield was recorded in SKUA-7A × RL-3. The marker-based assay using Rf3 and Rf4 linked markers was also carried out to validate the genes responsible for fertility restoration. The genotypes RL-1, RL-2, RL-5, RL-9, RL-11 and RL-12 showed polymorphism with all the markers. Sixteen hybrids showed fertility restoration behaviour with all the markers as well as under the field condition. Among these, 8 fall in the class of top 10 heterotic hybrids in the present study. The heterotic hybrids were evaluated for grain and cooking quality features. Out of the top 10 heterotic hybrids evaluated, nine were of medium to short grain types and only one SKUA-19A × RL-5 was of long grain type. Heterotic hybrids along with their parents showed intermediate amylose content which is preferable. [ABSTRACT FROM AUTHOR]

Abstract: In the rice cytoplasmic-genetic male sterility (CMS) system, the combination of a CMS line, maintainer line and restorer line carrying the restorer gene to restore fertility, is indispensable for the development of hybrids. However, the process of screening for the trait of fertility restoration is laborious and time-consuming. In the present study, we analyzed the nucleotide sequence of the Rf4 gene, which is the major locus controlling fertility restoration, to identify allele-specific variation. A single nucleotide polymorphism (SNP) A/C at +474 in the coding sequence (CDS) was found to be capable of strictly distinguishing groups of alleles Rf4 (A) and rf4 (C). Using KASP genotyping, this valuable SNP was converted to an allele-specific PCR marker. We evaluated and validated the marker among three-line parents with different backgrounds, and the results revealed a complete correlation between SNP alleles and the fertility restoration phenotype. Molecular screening was subsequently carried out for the presence of alleles of Rf4 and Rf3 among 328 diverse rice cultivars with worldwide distribution. The results demonstrate that this SNP marker could be the optimal choice for the molecular identification of potential restorers.

This study was carried out to improve the RPHR-1005, a stable restorer line of the popular medium slender grain type rice hybrid, DRRH-3 for bacterial blight (BB) and blast resistance through marker-assisted backcross breeding (MABB). Two major BB resistance genes, Xa21 and Xa33 and a major blast resistance gene, Pi2 were transferred to RPHR-1005 as two individual crosses. Foreground selection for Xa21, Xa33, Pi2, Rf3 and Rf4 was done by using gene-specific functional markers, while 59 simple sequence repeat (SSR) markers polymorphic between the donors and recipient parents were used to select the best plant possessing target resistance genes at each backcross generation. Backcrossing was continued till BC F and a promising homozygous backcross derived line possessing Xa21 + Pi2 and another possessing Xa33 were intercrossed to stack the target resistance genes into the genetic background of RPHR-1005. At ICF , 10 promising lines possessing three resistance genes in homozygous condition along with fine-grain type, complete fertility restoration, better panicle exertion and taller plant type (compared to RPHR-1005) were identified. [ABSTRACT FROM AUTHOR]

Two major nuclear genes, Rf3 and Rf4, are known to be associated with fertility restoration of wild-abortive cytoplasmic male sterility (WA-CMS) in rice. In the present study, through a comparative sequence analysis of the reported putative candidate genes, viz. PPR9-782-(M,I) and PPR762 (for Rf4) and SF21 (for Rf3), among restorer and maintainer lines of rice, we identified significant polymorphism between the two lines and developed a set of PCR-based codominant markers, which could distinguish maintainers from restorers. Among the five markers developed targeting the polymorphisms in PPR9-782-(M,I), the marker RMS-PPR9-1 was observed to show clear polymorphism between the restorer (n = 120) and maintainer lines (n = 44) analyzed. Another codominant marker, named RMS-PPR762 targeting PPR762, displayed a lower efficiency in identification of restorers and maintainers, indicating that PPR9-782-(M,I) is indeed the candidate gene for Rf4. With respect to Rf3, a codominant marker, named RMS-SF21-5 developed targeting SF21, displayed significantly lower efficiency in identification of restorers and non-restorers as compared to the Rf4-specific markers. Validation of these markers in a F2 mapping population segregating for fertility restoration indicated that Rf4 has a major influence on fertility restoration and Rf3 is a minor gene. Further, the functional marker RMS-PPR9-1 was observed to be very useful in identification of impurities in a seed lot of the popular hybrid, DRRH3. Interestingly, when RMS-PPR9-1 and RMS-SF21-5 were considered in conjunction with analysis, near-complete, marker–trait co-segregation was observed, indicating that deployment of the candidate gene-specific markers both Rf4 and Rf3, together, can be helpful in accurate identification of fertility restorer lines and can facilitate targeted transfer of the two restorer genes into elite varieties through marker-assisted breeding. [ABSTRACT FROM AUTHOR]

The elite Indian rice hybrid, DRRH3 is highly susceptible to two major diseases, bacterial blight (BB) and blast, which limit its productivity significantly. In the present study, we have introgressed two major genes, viz., Xa21 and Pi54 conferring resistance against BB and blast, respectively into RPHR-1005, the male parent of DRRH3 through marker-assisted backcross breeding (MABB) and analyzed the backcross derived plants for their resistance against BB and blast. RPBio Patho-2 was used as a donor for both the resistance genes. Gene-specific markers were used for the foreground selection of Xa21 and Pi54 at each stage of backcrossing and markers specific for the major fertility restorer genes, Rf3 and Rf4 were used only at BCF generation for foreground selection . Background selection was done using 62 polymorphic SSR markers and marker-assisted backcrossing was continued till BC generation. At BCF, through intensive phenotype-based selections 15 promising lines (ABLs) possessing high level of resistance against BB and blast, high yield, fine-grain type, complete fertility restoration along with better panicle exsertion and taller plant type as compared to RPHR-1005 were identified and test crossed with APMS 6 A, the female parent of DRRH3. The newly derived hybrids (i.e. improved versions of DRRH3) were observed to possess high level of resistance against BB and blast along with medium-slender grain type and yield level better than or equivalent to that of DRRH3. Our study exemplifies the utility of MABB for targeted improvement of multiple traits in hybrid rice. [ABSTRACT FROM AUTHOR]

RPHR-1005, the stable restorer line of the popular medium slender (MS) grain type rice hybrid, DRRH-3 was improved in this study for resistance against bacterial blight (BB) and blast diseases through marker-assisted backcross breeding (MABB). In this study, four major resistance genes (i.e., Xa21 and Xa33 for BB resistance and Pi2 and Pi54 for blast resistance) have been transferred to RPHR-1005 using RPBio Patho-1 (possessing Xa21 + Pi2), RPBio Patho-2 (possessing Xa21 + Pi54) and FBR1-15EM (possessing Xa33) as the donors. Foreground selection was carried out using PCR-based molecular markers specific for the target resistance genes and the major fertility restorer genes, Rf3 and Rf4, while background selection was carried out using a set of parental polymorphic rice SSR markers and backcrossing was continued uptoBC2 generation. At BC2F2, plants possessing the gene combination- Xa21 + Pi2, Xa21 + Pi54 and Xa33 in homozygous condition and with >92% recovery of the recurrent parent genome (RPG) were identified and intercrossed to combine all the four resistance genes. Twenty-two homozygous, pyramid lines of RPHR-1005 comprising of three single-gene containing lines, six 2-gene containing lines, eight 3-gene containing lines, and five 4-gene containing lines were identified among the double intercross lines at F3 generation (DICF3). They were then evaluated for their resistance against BB and blast, fertility restoration ability and for key agro-morphological traits. While single gene containing lines were resistant to either BB or blast, the 2-gene, 3-gene, and 4-gene pyramid lines showed good level of resistance against both and/or either of the two diseases. Most of the 2-gene, 3-gene, and 4-gene containing pyramid lines showed yield levels and other key agro-morphological and grain quality traits comparable to the original recurrent parent and showed complete fertility restoration ability, with a few showing higher yield as compared to RPHR-1005. Further, the experimental hybrids derived by crossing the gene-pyramid lines of RPHR-1005 with APMS6A (the female parent of DRRH-3), showed heterosis levels equivalent to or higher than DRRH-3. The results of present study exemplify the utility of MABB for targeted improvement of multiple traits in hybrid rice. [ABSTRACT FROM AUTHOR]

Twenty one premium grain quality genotypes of rice were used for identification of restorers and maintainers for 3 WA cytoplasmic male sterile lines. Six genotypes (Sanwal Basmati, Pusa Sugandh-2, Pusa Sugandh-3, Pusa Sugandh-5, Pusa 2517-2-51-1 and HUR-JM-59221) were found to exhibit stable restorer behaviour for all the 3 CMS lines (IR-58025A, IR-68897A and Pusa 6A). Moreover, Pusa-44 was found to exhibit the stable fertility restoring ability for IR-58025A, whereas Pusa Basmati-1121 revealed the stable restoring potential for IR-68897A. Three pollen parents (Pusa Basmati-1, Pusa-1460 and HUR-LP-191123) exhibited stable maintainer behaviour for all the 3 CMS lines. In addition, 3 genotypes i.e., HURPB-1M-98, HURPB-1S-97 and TBD-2-1 were found to show the stable maintainer behaviour for CMS line IR-58025A. The male parents showing stable fertility restoring behaviour along with the female parents and hybrids were evaluated with the help of molecular markers to establish the presence of fertility restoring genes. The primers RM171 and RM6100 linked to Rf4 gene revealed distinguishing banding pattern between CMS lines and fertility restorers, no such polymorphism was observed for the primers RM315 and RM443 linked to Rf3 gene. The genotypes HUR-JM-59221 and Pusa-44 revealed slightly different banding pattern for the primer RM171 than rest of the pollen parents. [ABSTRACT FROM AUTHOR]

The inheritance of the fertility restoration in cms-C in maize is very complex, and there are many contradictory results in the literature. There is an indication that there are different parallel or duplicated systems some of which include at least three complementary dominant genes - Rf4, Rf5, Rf6. Also, there is an indication of the duplication of the gene Rf4, which was tested in this work. We used three lines of different genetic constitution regarding Rf genes (B-73Bbms, L-70/9 and B-73cmsC-RfC) and produced different genotypes, combinations of genotypes and generations between them for the screening of fertility/sterility. The results of this work are negative, i.e. we did not prove the duplication of the Rf4 gene. This may be because of the relatively small number of lines that we have used (only three), or because there really is no duplication of the investigated gene. .

We have established marker-aided selection strategies for the two major Rf genes ( Rf3 and Rf4) governing fertility restoration of␣cytoplasmic-genetic male sterility (CMS) in rice. Polymorphisms between restorer and non-restorer␣lines were observed using RG140/ PvuII for Rf3 located on chromosome 1 and S10019 /BstUI for Rf4 located on chromosome 10 . DNA polymorphisms associated with these two loci in restorer lines of wild abortive (WA), Dissi, and Gambiaca cytoplasm are conserved, suggesting that similar biological processes control pollen fertility in this diverse cytoplasm. Because of their close linkage to Rf genes and distinct banding patterns, STS markers RG140/ PvuII and S10019/ BstUI are well suited for marker-aided selection, enhanced backcross procedures, and pyramiding of Rf genes in agronomically superior non-restorer lines. The combined use of markers associated with these two loci improved the efficiency of screening for putative restorer lines from a set of elite lines. Positional analyses of Rf4 and the inheritance pattern of the polymorphism in S10019/ BstUI suggest that Rf4, governing fertility restoration in WA-CMS in rice, is likely to be the same gene governing fertility restoration in BT- and HL-CMS that has a gametophytic effect, which explains why 100% pollen fertility in hybrids is impossible to attain. [ABSTRACT FROM AUTHOR]

The wild abortive cytoplasmic male sterility (CMS-WA) system, an ideal type of sporophytic CMS in indica rice, is used for the large-scale commercial production of hybrid rice. Searching for restorer genes is a good approach when phenotyping is very time-consuming and requires the determination of spikelet sterility in testcross progeny. To establish more precisely the genetical and physical maps of the Rf4 gene, high-resolution mapping of this locus was carried out using simple sequence repeat (SSR) markers and newly designed markers in a F2 population. The genetic linkage analysis indicated that five SSR markers (RM6737, RM304, RM171, RM5841 and RM228) on the long arm of chromosome 10 were linked with the Rf4 gene. Rf4 was flanked by two SSR markers RM171 and RM6737 at distances of 3.2 and 1.6 cM, respectively. Also, within the region between Rf4 gene and RM171, a newly designed primer pair, AB443, produced two sterile-specific markers, AB443-400 and AB443-500, 0.5 and 1.03 cM from the gene. The flanking markers identified give promise for their application in molecular marker-assisted selection (MAS) and they are also suitable for starting chromosome walking to clone Rf4 gene in the near future. [ABSTRACT FROM AUTHOR]

Peptidyl­tRNA dissociation from ribosomes is an energetically costly but apparently inevitable process that accompanies normal protein synthesis. The drop-off products of these events are hydrolysed by peptidyl-tRNA hydrolase. Mutant selections have been made to identify genes involved in the drop-off of peptidyl-tRNA, using a thermosensitive peptidyl­tRNA hydrolase mutant in Escherichia coli. Transposon insertions upstream of the frr gene, which encodes RF4 (ribosome release or recycling factor), restored growth to this mutant. The insertions impaired expression of the frr gene. Mutations inactivating prfC, encoding RF3 (release factor 3), displayed a similar phenotype. Conversely, production of RF4 from a plasmid increased the thermosensitivity of the peptidyl­tRNA hydrolase mutant. In vitro measurements of peptidyl­tRNA release from ribosomes paused at stop signals or sense codons confirmed that RF3 and RF4 were able to stimulate peptidyl­tRNA release from ribosomes, and showed that this action of RF4 required the presence of translocation factor EF2, known to be needed for the function of RF4 in ribosome recycling. When present together, the three factors were able to stimulate release up to 12-fold. It is suggested that RF4 may displace peptidyl­tRNA from the ribosome in a manner related to its proposed function in removing deacylated tRNA during ribosome recycling. [ABSTRACT FROM AUTHOR]

RPHR-1005, the stable restorer line of the popular medium slender (MS) grain type rice hybrid, DRRH-3 was improved in this study for resistance against bacterial blight (BB) and blast diseases through marker-assisted backcross breeding (MABB). In this study, four major resistance genes (i.e., Xa21 and Xa33 for BB resistance and Pi2 and Pi54 for blast resistance) have been transferred to RPHR-1005 using RPBio Patho-1 (possessing Xa21 + Pi2), RPBio Patho-2 (possessing Xa21 + Pi54) and FBR1-15EM (possessing Xa33) as the donors. Foreground selection was carried out using PCR-based molecular markers specific for the target resistance genes and the major fertility restorer genes, Rf3 and Rf4, while background selection was carried out using a set of parental polymorphic rice SSR markers and backcrossing was continued uptoBC2 generation. At BC2F2, plants possessing the gene combination- Xa21 + Pi2, Xa21 + Pi54 and Xa33 in homozygous condition and with >92% recovery of the recurrent parent genome (RPG) were identified and intercrossed to combine all the four resistance genes. Twenty-two homozygous, pyramid lines of RPHR-1005 comprising of three single-gene containing lines, six 2-gene containing lines, eight 3-gene containing lines, and five 4-gene containing lines were identified among the double intercross lines at F3 generation (DICF3). They were then evaluated for their resistance against BB and blast, fertility restoration ability and for key agro-morphological traits. While single gene containing lines were resistant to either BB or blast, the 2-gene, 3-gene, and 4-gene pyramid lines showed good level of resistance against both and/or either of the two diseases. Most of the 2-gene, 3-gene, and 4-gene containing pyramid lines showed yield levels and other key agro-morphological and grain quality traits comparable to the original recurrent parent and showed complete fertility restoration ability, with a few showing higher yield as compared to RPHR-1005. Further, the experimental hybrids derived by crossing the gene-pyramid lines of RPHR-1005 with APMS6A (the female parent of DRRH-3), showed heterosis levels equivalent to or higher than DRRH-3. The results of present study exemplify the utility of MABB for targeted improvement of multiple traits in hybrid rice.