**2. Molecular marker**

Rediscovery Mendelian theory in the early part of 20th century, reveal that inheritance (genes) linked together in chromosome. Molecular marker defines as individual genes flanking within a defined close interval. It can be found at a specific location of the genome and linked with inheritance of a trait or gene [13]. Thottappilly et al. [14] refers molecular marker as polymorphism identified between species including proteins and nucleic acids. Collard et al. [15] had divided molecular marker into 3 major groups which are morphological marker, biochemical marker and DNA marker (**Figure 1**). However, Xu [16] had categorized markers into two

**103**

**Table 2.**

*Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.)*

and deletions) or errors in replication of tandemly repeated DNA [18].

Advantages Readily available Highly reproducible Usually require only simple

The most direct measure of

Subjected to environmental

equipment

phenotype

influences

Application Conventional plant breeding program

*Comparison between classical markers.*

laboratory

multiple loci SSR Reliable, powerful and easy

SNP Multiplexing large number of markers

used

*Comparison of different PCR based marker.*

Simple, rapid and lowest

Single primer generates

Transferable between populations

Degraded DNA can be

RAPD A small amount of DNA required

cost

Limited in number

Disadvantages Requires expertise on crop or species

broad groups based on detecting method; (i) classical marker and (ii) DNA marker. Classical marker was initiated with the mutations in the genomic loci controlling plant morphology [17] (**Table 1**). Advent in polymorphism detecting methods such as Polymerase Chain Reaction, Southern Blotting and Sequencing had invented the development of DNA markers (**Table 2**). Variation were detected at DNA level such as nucleotides changes ie; substitution (point) mutation, rearrangement (insertion

**Morphological marker Biochemical marker**

Applicable for measure a wide range of population genetic parameter

Limited biochemical assay for detection

Hybrid fixation

Saturation mapping

Linkage and QTL mapping Marker-assisted selection Hybrids fixation

Map-based cloning

Phenotype-based analysis

Genetic diversity Map construction

**Advantages Disadvantages Application**

Genetic diversity Easy to conduct across

Irreproducibility

establishment

electrophoresis

Marker has less allele

AFLP Multiple loci Required several DNA Saturation mapping

Required a larger amount of DNA

Tedious and costly for initial

Required polyacrylamide gel

Not transferable across laboratory Genetic diversity

Genetic diversity Complicated

Costly to assay Fine mapping

RFLP Reliable and rigid Harmful and tedious Map construction

Inexpensive

*DOI: http://dx.doi.org/10.5772/intechopen.93126*

**Types of marker**

**Table 1.**

**PCR based markers**

**Figure 1.** *Different types of molecular marker available.*

#### *Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.) DOI: http://dx.doi.org/10.5772/intechopen.93126*

broad groups based on detecting method; (i) classical marker and (ii) DNA marker. Classical marker was initiated with the mutations in the genomic loci controlling plant morphology [17] (**Table 1**). Advent in polymorphism detecting methods such as Polymerase Chain Reaction, Southern Blotting and Sequencing had invented the development of DNA markers (**Table 2**). Variation were detected at DNA level such as nucleotides changes ie; substitution (point) mutation, rearrangement (insertion and deletions) or errors in replication of tandemly repeated DNA [18].


#### **Table 1.**

*Recent Advances in Rice Research*

mapping and gene pyramiding approaches.

**2. Molecular marker**

Several amendments can be implemented to overcome these limitations such as breeding high yielding varieties, durable resistance to diseases and tolerance to abiotic stresses [5]. Rice yield potential can be enhanced by using numerous approaches such as conventional hybridization, ideotype breeding, heterosis breeding, wide hybridization and molecular breeding [6]. In molecular rice breeding, there are two main strategies can be used by applying the biotechnology concept which are MAS and Genetically Engineering. MAS like genetic engineering is a technique used to introgress targeted gene however its offers tools for specific selection for further breeding in existing plant material. The field of plant genetics and breeding has irreversibly changed with the development of DNA (or molecular markers) [7]. Since past two decade, revolutionized in molecular marker to enhance the effectiveness in breeding and to significantly shorten the development time of varieties bring about the concept of molecular marker assisted selection (MAS) as an efficient plant selection [8–10]. A genetic marker is any noticeable character or otherwise assayable phenotype, for which alleles at individual loci segregate in a Mendelian manner. The genetic markers covered include (1) morphological markers (2) biochemical markers (alloenzymes and other protein markers) and (3) molecular markers (based on DNA-DNA hybridization) [11, 12]. In this chapter, we present the latest DNA-based markers in a few studies related to abiotic and biotic stress genes using the MAS application in rice breeding programs. These markers have proven significantly useful from different findings published in screening, fine

Rediscovery Mendelian theory in the early part of 20th century, reveal that inheritance (genes) linked together in chromosome. Molecular marker defines as individual genes flanking within a defined close interval. It can be found at a specific location of the genome and linked with inheritance of a trait or gene [13]. Thottappilly et al. [14] refers molecular marker as polymorphism identified between species including proteins and nucleic acids. Collard et al. [15] had divided molecular marker into 3 major groups which are morphological marker, biochemical marker and DNA marker (**Figure 1**). However, Xu [16] had categorized markers into two

**102**

**Figure 1.**

*Different types of molecular marker available.*

*Comparison between classical markers.*


#### **Table 2.** *Comparison of different PCR based marker.*

Although there are many characteristics determine the suitable markers to be considered in MAS development; there are five main characteristics need to be considered [13, 19–22].


### **3. Successful application of molecular marker in rice**

Recently, MAB has become a well-known approach develop a superior genotype in rice. The aims of the breeding programs were to develop a rice variety with high yield potential, resistant to diseases and insects, tolerance to adverse environments, and acceptable grain quality. Since the first successful experiment stated for rice by Chen et al. [24] with introducing resistance to bacterial blight (BB) disease into Chinese hybrid parents. Till date, many varieties had been developed thru the application of molecular marker with backcrossing method. Varieties developed were improved from the wild type by incorporate desirable gene depending for biotic and abiotic stresses. Recently, in a few years there is a trend in producing more rice varieties that resistant in abiotic stress such as drought tolerance, submergence tolerance and salt tolerance. This demand comes from the periodic natural disaster in many regions of the rice producing country specifically in Asian country [25]. Resistant varieties against pathogenic disease were continuously developed. This scenario can be seen in blast disease where there are increasing number of resistant varieties were developed due to instability of the fungus pathogen.

Numerous molecular markers have been utilized in marker-assisted breeding however microsatellites are the preferable markers for plant breeding applications. The characteristic as codominant markers which inherited in a Mendelian fashion have emerged as a best choice compare to other markers. Additionally, microsatellite identify as preferable markers used in plant breeding programs due to ubiquitous, high polymorphism rates and wide range of distribution throughout the genome [26, 27]. Other markers possess major drawbacks including Restriction fragment length polymorphism (RFLP) markers which are not integrated with high-throughput technique; Random amplification of polymorphic DNA (RAPD) assays which are often not reproducible or immobile. AFLP analysis is not straightforward and commonly produces multiple fragments with the use of large genomic templates [28]. Application of molecular marker in rice breeding was demonstrated in **Table 3** to develop superior varieties in rice against abiotic and biotic stresses.

**105**

**Stress** Abiotic

Deep roots

Quality Root traits and aroma

Submergence tolerance

*Sub1* QTL *Sub1* QTL

*Sub1*QTL *Sub1* QTL

*Sub*1

*Sub1* *Sub1* *Sub1* *Saltol* *Saltol* QTL *Saltol* QTL *Saltol* QTL

*Hd2*

Early maturation

High yield and drought tolerance

*qDTY12*

*qDTY1.1, qDTY2.2, qDTY3.1, qDTY3.2, qDTY6.1*, and

*qDTY1.1 + qDTY2.1 + qDTY3.1 + qDTY11.1 and two QTLs* 

*qDTY1.1 + qDTY11.1*

QTL (*Hd1, Hd4, Hd5* and *Hd6*)

Heading time Phosphorus tolerance

Drought tolerance

*Pup1* QTL MQTL1.1

QTL

Salt tolerance

**Traits**

**Gene/QTLs** QTLs on chromosomes 1, 2, 7 and 9

waxy QTLs on chromosomes 2, 7, 8, 9 and 11

**Foreground and background selection**

RFLP and SSR

RFLP RFLP and SSR Phenotyping and SSR

SSR SSR SSR SSR SSR SSR SSR SSR SSR SSR SNP SSR SSR SSR RFLP, STS, SSR,

RFLP, STS, SSR,

[47]

CAPS, dCAPS

SSR SSR SSR SSR

[48]

[42]

[49]

[50]

CAPS, dCAPS

SSR SSR SSR SSR

SSR

[46]

SSR SSR STS SSR SSR SSR SSR SSR SSR SSR SSR SSR SSR SSR

SSR AFLP RFLP and SSR

[29] [30] [31] [32] [33] [34] [35] [36] [37] [38] [39] [40]

[41]

[42]

[43]

[44]

[45]

**Reference**

*Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.)*

*DOI: http://dx.doi.org/10.5772/intechopen.93126*


*Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.) DOI: http://dx.doi.org/10.5772/intechopen.93126*

*Recent Advances in Rice Research*

considered [13, 19–22].

process.

software are significant for this purpose.

Although there are many characteristics determine the suitable markers to be considered in MAS development; there are five main characteristics need to be

1. **Reliability:** Attribute of the desirable marker need to close proximity with the target loci, preferably less than 5 cm genetic distance or one marker every 10 cm [23].

2.**Repeatability and reproducible:** In plant breeding, a lot number of plants are usually screened for desirable marker pattern and result is required instantly. Therefore, level of simplicity in terms of time required and quick detection method are highly desirable for rapid selection. Statistical and bioinformatics

3.**Cost effective:** Marker assay should be not expensive feasible for selection

4.**Level of polymorphism:** Marker should discriminate between two similar

5.**DNA quality and quantity:** A few markers require a complex DNA extraction technique and it too laboratories. In addition, it adds cost to the process.

Recently, MAB has become a well-known approach develop a superior genotype in rice. The aims of the breeding programs were to develop a rice variety with high yield potential, resistant to diseases and insects, tolerance to adverse environments, and acceptable grain quality. Since the first successful experiment stated for rice by Chen et al. [24] with introducing resistance to bacterial blight (BB) disease into Chinese hybrid parents. Till date, many varieties had been developed thru the application of molecular marker with backcrossing method. Varieties developed were improved from the wild type by incorporate desirable gene depending for biotic and abiotic stresses. Recently, in a few years there is a trend in producing more rice varieties that resistant in abiotic stress such as drought tolerance, submergence tolerance and salt tolerance. This demand comes from the periodic natural disaster in many regions of the rice producing country specifically in Asian country [25]. Resistant varieties against pathogenic disease were continuously developed. This scenario can be seen in blast disease where there are increasing number of resistant

accessions of species especially in parent breeding material.

**3. Successful application of molecular marker in rice**

varieties were developed due to instability of the fungus pathogen.

Numerous molecular markers have been utilized in marker-assisted breeding however microsatellites are the preferable markers for plant breeding applications. The characteristic as codominant markers which inherited in a Mendelian fashion have emerged as a best choice compare to other markers. Additionally, microsatellite identify as preferable markers used in plant breeding programs due to ubiquitous, high polymorphism rates and wide range of distribution throughout the genome [26, 27]. Other markers possess major drawbacks including Restriction fragment length polymorphism (RFLP) markers which are not integrated with high-throughput technique; Random amplification of polymorphic DNA (RAPD) assays which are often not reproducible or immobile. AFLP analysis is not straightforward and commonly produces multiple fragments with the use of large genomic templates [28]. Application of molecular marker in rice breeding was demonstrated in **Table 3** to develop superior varieties in rice against abiotic and biotic stresses.

**104**


**107**

**Stress**

**Traits** Submergence tolerance, disease resistance, quality Thermo-sensitive genic male sterile (TGMS) and blast resistance

TGMS and *Pi* gene

Brown planthopper resistance

Gall midge resistance

Rice stripe resistance

**Table 3.** *Application of molecular marker in MAB in rice.*

*Bph14* and *Bph15*

*Bph18*

*Gm8* *Stv-bi*

**Gene/QTLs** *Subchr9* QTL, *xa21*, *Bph* and blast QTLs and quality loci

SSR and STS

SNP InDEL

SSR SSR SSR

InDEL

SSR SSR SSR

[74]

SNP

[73]

**Foreground and background selection**

Not performed

[34]

**Reference**

*Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.)*

*DOI: http://dx.doi.org/10.5772/intechopen.93126*

[75] [76] [77]

#### *Recent Advances in Rice Research*


*Recent Advances in Rice Research*

**106**

**Stress** Biotic

Bacterial blight

**Traits**

**Gene/QTLs** (qDTY1.1, qDTY2.1)

QTL *xa21* *xa21* *xa5, xa13* and *xa21*

*xa5, xa13* and *xa21*

*xa33* *xa34(t)* *xa35(t)*

*xa38* *xa21*

*xa5* *xa13, xa21* *xa5* and *xa13* *xa13* and *xa21* *xa13+ xa21* and *xa5 + xa21*

*xa13 and xa21* *Pi1, Pi2, Pi33* and *Pi54*

*Pi* genes, *xa5*

*Pi54* and *Pi5*

*Pi1* and *Pi2*

*Pi1* *Pikh and pi7(t)*

*QTLs on chromosomes 1, 2, 11 and 12*

Blast resistance

**Foreground and background selection**

SSR SSR STS STS STS, CAPS

STS SSR SSR SSR SSR STS CAPS

STS CAPS CAPS

SSR SSR SSR SSR SSR SSR SSR SSR SSR

SSR SSR RFLP AFLP Not performed Not performed

SSR SSR SSR SSR STS CAPS

SSR STS STS SSR SSR SSR SSR SSR SSR ISSR

SSR SSR

[51]

[52]

[24]

[53]

[54]

[55]

[56]

[57]

[58]

[59]

[60]

[61]

[62]

[63]

[62, 64]

[65]

[66]

[67]

[68]

[69]

[70]

[71]

[9]

[72]

**Reference**

**Table 3.** *Application of molecular marker in MAB in rice.*

#### *Current Applicable DNA Markers for Marker Assisted Breeding in Rice (*Oryza sativa *L.) DOI: http://dx.doi.org/10.5772/intechopen.93126*
