**5. Cytochrome P450 monooxygenases (P450S) and evolved herbicide resistance**

Sanger sequencing [13, 124]. In many studies, these approaches were used to uncover TSR in

Recognizing alleles of non-target-site-based resistance requires the identification of alleles specific for resistant genotypes while contrasted with sensitive genotypes, and to eliminate "false positives." These alleles are diverse in both genotypes (resistant and susceptible), however, do not involve in non-target-site-based resistance. In plant genomes, there are numerous alleles reported to be associated with non-target-site-based herbicide resistance. Alleles associated with quantitative characteristics are mostly identifies using genetic marker approaches (quantitative trait loci (QTL) mapping) [132]. QTL mapping is intricate, time consuming, and not easily applied to natural or field populations of nonmodel organisms such as weeds [133]. Another approach utilized for identification of alleles dedicating quantitative properties is to interrupt or imitate the phenotype of interest through genetic transformation [134]. Owing to the recent technical and scientific "omics" revolution, the genetic basis of quantitative characteristics, such as NTSR, can be explained even in nondemonstrate species, for example weeds.

To accomplish this objective, three stages ought to be completed (**Figure 2**).

**Figure 2.** Three-step procedure to identify NTSR alleles [8].

weeds [125–131].

**4.2. Non-target-site resistance**

112 Herbicide Resistance in Weeds and Crops

P450s are one of the largest super families of enzymes. Plants have the most noteworthy number of P450 genes. It is well demonstrated that tolerance to several modes of action to some herbicides is associated with Cyt P450 mediated enhanced metabolism. Cytochrome P450s in plants synthesize sterols, fatty acid derivatives, and hormones and in some cases, these are involved in plant secondary metabolism. Usually, plant cytochrome P450s are limited in the endoplasmic reticulum (in a few cases to plastid membranes). When a vast variety in plant metabolism catalyzes by cytochrome P450s, the role of cytochrome P450s is often hydroxylation or dealkylation in the herbicide alteration. Several plant cytochrome P450s will metabolize some herbicides to further inactivated productions so that their phytotoxicity will decrease or alter. Mostly, this process occurs by conjugation to glucose and subsequent transport into the vacuole. [36, 46, 135]. As crops can P450 metabolize many different herbicides, their use on large weed populations is a strong selection pressure for weed individuals possessing the same ability. Indeed, in weeds, P450-based herbicide resistance is a very threatening resistance mechanism because P450 enzymes can simultaneously metabolize herbicides of different modes of action, potentially including never-used herbicides [136, 137]. Subsequently, *in vivo* studies on herbicide metabolism and P450 inhibitors in resistant biotypes showed that P450s catalyzed enhanced rates of metabolism of several herbicides [138–140]. In addition to *L. rigidum* and *A. myosuroides*, the evolution of resistance due to P450-catalyzed enhanced rates of herbicide metabolism has been demonstrated in resistant biotypes of a further nine weed species [57, 141]. Cytochrome P450s can metabolize low dose of diclofop-methyl in some susceptible biotypes of *L. rigidum* and therefore the weeds were treated at a dose bringing around 50% mortality. Some survivors were grown for producing seeds to create the next generation, and in this direction, the selection was repeated. Nontarget-site resistance was progress of high level in lonely three generations [142]. Importantly, there was concomitant evolution of cross-resistance to other P450-metabolizable herbicides of different modes of action [85, 88]. In *L. rigidum*, evolved P450-based herbicide resistance can be correlated with a fitness cost [143, 144]. Up to now, slight data are obtained through biochemical investigation to determine P450-based herbicide resistance in some evolved resistant weed species. Cytochrome P450 microsomes which degrade herbicides have not been successfully isolated from resistant *A. myosuroides* and *L. rigidum*, while they have been successfully isolated from resistant *E. phyllopogon* [145]. Isolated 16 P450 genes from a resistant *L. rigidum* biotype were not imputed to herbicide metabolism [76, 146]. One of three fulllength P450s, CYP71R4, were gained from resistant *L. rigidum* biotypes metabolized a PSII herbicide, while expressed in yeast [147]. P450 proteins can share as little as 16% amino acid identity, and there are more than 2000 plant P450 sequences in the P450 database. Reportages about P450-based evolved herbicide resistance in grass weed species has often been more than dicot species, whereas dicot weed species have less P450 genes toward grass weed species. This fact reflects the attitude of some investigate to test only for target-site-based resistance [17].
