Second, we image individuals on apple juice agar supplemented with a dye for contrast enhancement over background. ![]() First, we use a simple camera setup that provides consistency from one experiment to the next while being accessible to any lab. This system has several advantages over other larvae tracking setups. Herein, we have modified the optimal settings of the wrMTrck program for the detection and analysis of Drosophila 元 larvae. elegans crawling on agar plates or swimming in liquid ( Husson et al., 2012). Our lab has modified a freeware version of the ImageJ Plugin wrMTrck (ImageJ: ) and wrMTrck: ), originally designed to track the movement and speed of C. While these automatic phenotyping programs are useful for large scale projects in which analysis of multiple genotypes is needed, simpler tracking systems are required for basic locomotor analysis. A computerized, single-animal tracking system, called MaggotTracker, was also devised to measure and analyze over 20 different parameters of larval traits, including shape, size, and peristalsis movements (Aleman-Meza, et al., 2015). ![]() Risse et al., designed an imaging technique based upon frustrated total internal reflection (FTIR) to generate high resolution movies that permit forward genetic screens ( Risse et al., 2013). Other groups have generated technological imaging-based approaches that can be used for the rapid, high throughput analysis of various larval locomotor behaviors. One approach to circumvent this problem is to feed larvae colored dyes to enhance contrast followed by movement analysis using the commercially available Matlab software ( Khurana et al., 2010). Unfortunately, the semi-translucent nature of the Drosophila larval body results in low contrast between the organism and background on standard apple juice media. Computer-based tracking programs have been developed for Drosophila and other organisms that rely on visualizing pixel intensity differences between the specimen and background ( Sinadinos et al., 2012, Jung et al., 2015, Gomez-Marin et al., 2012). However, this method can be time-consuming and the tracks of individual larvae are not recorded. A simple, yet effective method is manual tracking on a gridded surface, which counts the number of squares traversed by larvae in a specified time period ( Wang et al., 2015). ![]() There are numerous published assays for examining larval movement. Thus, phenotypic analysis is performed in the prior third larval instar (元) stage. For example, the muscle mutants tiggrin (tig) thin (tn) and muscle LIM protein (mlp84B) are all pupal lethal ( Bunch et al., 1998, LaBeau-DiMenna et al., 2012, Clark et al., 2007). Additionally, analysis of the larval stage is necessary if mutants do not survive to adulthood. ![]() Larval behavior can be reliably monitored to examine the effects of drugs or altered expression of human disease genes ( Pandey & Nichols, 2011, van der Voet et al., 2014). However, larval movement is relatively low speed, making it feasible to track both velocity and crawling patterns. The flight ability of Drosophila adults makes accurately recording speed and trajectories difficult. Both the larval and adult stages are routinely used in assays to monitor locomotion, feeding, circadian, and learning behaviors ( ). Drosophila melanogaster has emerged as a critical model organism to understand how altered behavior is linked to human disease phenotypes and aging.
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