*3.1.2 Marker-assisted selection (MAS)*

Marker-assisted selection is a method of breeding that makes use of molecular markers connected to particular qualities of interest. Breeders can selectively breed maize varieties with those markers, expediting the development of desired features, by discovering genetic markers linked to traits related to the generation of biofuels, such as high sugar or starch content [28].

#### *3.1.3 Genomic selection*

Genetic information about a person is used to predict that person's performance. Breeders can determine the genetic potential of various individuals for features linked to biofuels by studying the full genome of maize plants. The choice of parent lines for crosses can be influenced by this knowledge, improving the output of biofuel in succeeding generations [29].


#### **Table 1.**

*Different aspects of corn breeding for biofuel production.*

#### *3.1.4 Genetic engineering*

To improve features relevant to biofuel production in maize, certain genes can be inserted using genetic engineering techniques. For instance, genes that boost stress resistance or lower lignin content can be introduced into the genomes of maize plants. Breeders can introduce unique features that might not be present in the maize gene pool thanks to the exact alterations made possible by genetic engineering.

#### *3.1.5 Doubled haploid (DH) method*

Homozygous plants with all of their genes present in a single plant are created using the DH method and this strategy can hasten the creation of pure breeding lines with desired features for biofuels. The use of DH technology shortens the breeding process and enables the selection of superior lines based on characteristics including disease resistance, sugar content, and biomass production [26].

#### *3.1.6 High-throughput phenotyping*

To quickly and precisely evaluate a variety of plant features, high-throughput phenotyping uses automated methods. Breeders can choose and create better varieties by identifying individuals with superior biofuel-related features using high-throughput phenotyping systems to analyze vast populations of maize plants [27].

#### *3.1.7 Multi-trait selection*

Because the generation of biofuels is so complex, it's crucial to take several features into account at once. For which, breeders can apply multi-trait selection techniques that take into account traits like disease resistance, stress tolerance, and nutrient utilization efficiency in addition to biomass output and sugar or starch content. It guarantees a thorough improvement in maize varieties for the production of biofuels.

It's also imperative to note that various breeding techniques are frequently combined and integrated to increase their efficacy. The choice of breeding strategy is also influenced by the resources that are available, the breeding objectives, and the regulatory factors.
