Box plots of peak areas of individual waxes and total wax on the foliage of plants of semi-glossy SG-Syn and the waxy doubled haploid (DH) 2107.
Scatter plots with regression lines and percent coefficients of determination (R2) for peak areas from S1 progenies and mean peak areas of parents for the ratio of total fatty alcohols (FAs) over amounts of Hentriacontane (Hent) and 16-Hentriacontanone (H16) [FA·(Hent+H16)−1] (A) and mean peak areas for total wax (B).
Boxplot of fatty alcohol (FA)·(Hent+H16)−1 for the parental synthetic population (SG-Syn), low and high phenotypic selections, and cycle 1 of S1 family selection. Horizontal lines in boxes are the median. The letter X indicates the mean value. H16 = 16-Hentriacontanone; Hent = Hentriacontane.
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Epicuticular waxes on onion (Allium cepa) foliage offer protection against water loss, pathogens, and pests. Onion leaves with a light green color are referred to as “glossy” and have less total wax compared with that of darker blue-green (“waxy”) foliage. Leaves with intermediate amounts of wax can be visually classified as “semi-glossy.” Onion plants with glossy or semi-glossy foliage support fewer onion thrips (Thrips tabaci) and experience less feeding damage relative to that of plants with waxy foliage. The main goals of this study were to use gas chromatography–mass spectrometry (GCMS) to test the significance of parent–offspring regressions for waxes on leaves of semi-glossy onion and measure the response to selection for a specific wax profile. Plants from different sources of the semi-glossy phenotype were identified in field plots, intercrossed, and segregating families were produced. Complementation analyses revealed that different sources of the semi-glossy phenotype had the same genetic basis. A synthetic population (SG-Syn) was developed by intercrossing among semi-glossy progenies from the segregation analysis, and field-grown plants of SG-Syn were visually semi-glossy. Individual plants from SG-Syn were grown in a greenhouse, the amounts and types of leaf waxes were determined using GCMS, and plants were self-pollinated. Progenies were evaluated in the greenhouse to determine wax composition and amounts. Significant regression coefficients were observed for amounts of major waxes on leaves of S1 progenies and parents, indicating that wax profiles are heritable. Cycles of phenotypic and S1-family selection for the ratio of the amount of fatty alcohols over the amount of an alkane and ketone were completed. Significant responses to selection were observed, thus revealing that this wax profile is heritable. These results should be useful for the selection of unique epicuticular wax profiles with the goal of potentially developing thrips-resistant onion cultivars.
Epicuticular waxes accumulate on aerial surfaces of terrestrial plants to prevent water loss as well as protect against diseases and pests (Baker 1982). Epicuticular waxes are made of several different classes of lipids, including fatty acids and alcohols, ketones, aldehydes, and alkanes (Eigenbrode and Espelie 1995; Samuels et al. 2008). The composition of lipids that form epicuticular waxes differs among plant species (Baker 1982; Eigenbrode and Espelie 1995). In most plants, the synthesis of epicuticular waxes begins with the production of long-chain (>26 carbons) fatty acids that enter the acyl reduction or decarbonylation pathways (Millar et al. 1999; Samuels et al. 2008). In the acyl reduction pathway, fatty alcohols (FAs) with even numbers of carbons are produced. In the decarbonylation pathway, fatty acids lose one carbon to yield waxes with odd numbers of carbons.
Onion foliage can be visually classified based on relative amounts of epicuticular waxes. Onion leaves that have a light green color and accumulate less total wax can be classified as “glossy,” whereas leaves with darker blue–green foliage and higher amounts of waxes are referred to as “waxy.” Onions with intermediate amounts of waxes can be referred to as “semi-glossy” (figure of foliar phenotypes was presented by Munaiz et al. 2020). Khosa et al. (2020) reported that the primary waxes on onion (Allium cepa) foliage are the following: FAs 1-Heptadecanol [Heptol (C17)], 1-Hexacosanol [Hex (C26)], 1-Octacosanol [Oct1 (C28)], and 1-Triacontanol [Tri (C30)]; alkanes Heptacosane (Hept (C27)], Nonacosane [Non (C29)], and Hentriacontane [Hent (C31)]; ketone 16-Hentriacontanone [H16 (C31)]; and ether 1-Ethenyloxy-octadecane [Eth (C20)]. Heptol (C17) and Eth (C20) may be degradation products from longer-chain waxes or originate from different pathways. Previous studies reported that the total amount of wax and the relative proportion of H16 to total wax are associated with the visual appearance of leaves; onions with lower amounts of total wax and/or lower proportions of H16 relative to total wax have the glossy or semi-glossy phenotype, while onions with larger amounts of total wax and a higher proportion of H16 appear waxy (Damon et al. 2014; Munaiz et al. 2020).
Onion thrips (Thrips tabaci) are the most damaging insect pest of onion and cause large losses to both bulb and seed yields (Diaz-Montano et al. 2010; Fournier et al. 1995; Jones et al. 1934). Feeding by thrips larvae causes the greatest damage to onion leaves (Coudriet et al. 1979), and thrips can transmit Iris yellow spot virus, causing further losses (Gent et al. 2004; Pozzer et al. 1999). Control of thrips is difficult because of their high reproductive capacity, commonly occurring host plants, and overlapping generations. The insect tends to accumulate between the narrow leaves of onion, where sprays may not penetrate (Jones et al. 1934: Lall and Singh 1968). To control thrips, growers spray insecticides many times over the course of the growing season at substantial cost, and the insect has developed resistance to commonly used insecticides (Allen et al. 2005; Herron et al. 2008; Shelton et al. 2003, 2006). Development of onions with thrips resistance would allow growers to reduce sprays and potentially delay the development of pesticide resistance.
Epicuticular waxes affect interactions between plants and their insect pests (Eigenbrode and Espelie 1995). Onion plants with glossy leaves suffer less thrips damage relative to that of waxy plants (Boateng et al. 2014; Cramer et al. 2014; Damon et al. 2014; Diaz-Montano et al. 2010, 2012; Jones et al. 1934; Molenaar 1984). However, glossy onions have not been commercially grown because the relatively low amounts of leaf waxes are associated with susceptibility to powdery mildew (Mohan and Molenaar 2005) and damage by herbicides. Onions with semi-glossy foliage show less feeding damage by thrips and benefit from greater amounts of leaf waxes relative to glossy plants (Damon et al. 2014; Diaz-Montano et al. 2010; Munaiz et al. 2020). The semi-glossy foliage of onion inbred line ‘B5351’ is conditioned by two major quantitative trait loci (QTL) (Damon and Havey 2014). The first QTL is on chromosome 2 and primarily affects the amounts of the FAs; the second QTL is on chromosome 5 and is associated with the amounts of H16 and total wax. The visual phenotype (waxy vs. semi-glossy) mapped to the QTL on chromosome 5 (Damon and Havey 2014).
In this study, gas chromatography–mass spectrometry (GCMS) was used to measure variability for amounts and types of epicuticular waxes on leaves of semi-glossy onion accessions, different sources of the semi-glossy phenotype were intercrossed for a complementation test, and a synthetic semi-glossy population was created. The tested hypothesis was that a specific wax profile is heritable and can be altered by selection. The long-term objective of this research is the development of onion populations with unique epicuticular wax profiles to potentially reduce losses caused by onion thrips.
Onion seeds of plant introductions (PIs) listed in the USDA Germplasm Resource Information Network (GRIN-Global 2023) as having “low bloom” were obtained (Havey 2022). Seeds were planted in a field on the Kincaid Family Farm (Palmyra, WI, USA), and plants were grown under normal production conditions. Foliage was scored over the summer as glossy, semi-glossy, or waxy by visual comparisons with glossy (B9885) (Damon et al. 2014) and semi-glossy (B5351) (Damon et al. 2014) inbreds, and the waxy doubled haploid (DH) 2107 (Hyde et al. 2012). Bulbs of plants with semi-glossy foliage (that is, wax levels visually intermediate between glossy and waxy foliage) were harvested from inbred B5351 and PIs 255462, 264320, 264327, 321385, 546188, and 546192 (Table 1). After vernalization by storage at 4 to 7 °C for 6 months, pairs of bulbs were planted for crossing in a field at the University of Wisconsin Arlington Research Farm as follows: 264320 with B5351, 255462 with 264320, 264320 with 546192, 546192 with 546188, B5351 with 546192, 264327 with 264320, 321385 with 264320, and 546188 with 264327. Umbels of paired plants were covered with mesh cages and fly pupae (Calliphora spp.; Forked Tree Ranch, Bonners Ferry, ID, USA) were introduced for crossing (Havey 2018). Umbels were harvested separately from each plant. Seed from each umbel was cleaned and planted in a greenhouse on the University of Wisconsin–Madison campus. DNA was isolated using a mini-preparation (Macherey-Nagel Nucleospin 96 Plant II DNA kit; Bethlehem, PA, USA) from approximately 25 plants from each of the PIs and B5351 as well as individual progenies from crosses. DNA concentrations were determined spectrophotometrically (Nanodrop ND-1000; Thermo-Fisher, Waltham, MA, USA) and quality was assessed by electrophoresis through 1% agarose gels compared with uncut λ DNA. Twenty-five single nucleotide polymorphisms (SNPs) were genotyped (Duangjit et al. 2013) to identify hybrids from each of the crosses. Hybrids were transplanted to the field, bulbs were produced and vernalized, and plants were self-pollinated using flies as described above. Families from self-pollinations were grown in the field at Palmyra, WI, USA, and plants were visually scored according to leaf waxiness compared with adjacent plots of glossy and waxy onion. The synthetic population (SG-Syn) was developed by random mating using flies among 10 F2 progenies from each of eight families from the complementation test (Table 1), followed by another generation of random mating.
Field-produced bulbs of plants of SG-Syn and waxy DH2107 were harvested in September and vernalized at approximately 4 to 7 °C. In February of the following year, bulbs were planted in a soilless mix (Premier Tech, Riviere-du-Loup, Quebec, Canada) in classic 400 pots (4.5 L; Gemplers, Mt. Horeb, WI, USA) in a greenhouse at the University of Wisconsin–Madison campus. Plants were grown at 25 °C days and 20 °C nights with supplemental lighting at 14 h, watered as needed, and fertilized twice each week with 20N–8.7P–16.6K (Peter’s Professional 20–10–20; Everris, Dublin, OH, USA). At 8 and 9 weeks after bulb planting and before any pesticide sprays, leaves were sampled to measure epicuticular waxes. One fully expanded middle-aged leaf (oldest leaves and younger expanding leaves were avoided) was harvested separately from each of 72 plants of SG-Syn and six plants of DH2107. The leaf was held with forceps and cut with scissors, and an 8- to 10-cm leaf piece was placed in a glass tube (20 × 120 mm; Fischer Scientific, Waltham, MA, USA). The region of the leaf touched with forceps was discarded. After each sampling, forceps and scissors were dipped in chloroform and air-dried. After all samples were taken, 21 mL of high-performance liquid chromatography-grade (HPLC) chloroform (Sigma-Aldrich, St. Louis, MO, USA) was added to each tube, and forceps were used to submerge the leaf piece in chloroform for 1 min. After each sample, forceps were dipped in chloroform and air-dried. Leaf pieces were removed from the chloroform, placed in a separate tube, and dried in an oven at 80 °C for 4 d to allow measurement of dry weights. Then, 25 µL of docosane (1 mg·mL−1; Sigma-Aldrich) was added to each tube as an internal standard. Samples were prepared for GCMS as described below.
After leaf sampling, individual plants from SG-Syn were allowed to flower in the greenhouse and covered with mesh cages, and fly pupae were introduced for self-pollinations. Umbels were harvested from each plant and seed were cleaned. Seed from each of 44 S1 families and the original SG-Syn population were planted in the soilless mix (described) in each of three greenhouses in a randomized complete block design (greenhouses as blocks) under environmental conditions described previously except for supplemental lighting at 13 h. Two months after planting, one approximately 4-cm leaf segment was harvested from 10 plants from each S1 family, and the 10 leaf segments were combined into a single tube for each family. Leaf segments were harvested from 10 plants of SG-Syn and DH2107 and combined into separate tubes. One week later, a second sampling was completed from the same plants and leaf segments were combined in tubes as described above.
Ten plants from each of four S1 families with the highest FA·(Hent+H16)−1 ratios (mean: 1.10 ± 0.07 vs. 0.80 ± 0.25 for all S1 families) were moved in early January into a cold room at approximately 7 °C with 12-h fluorescent lighting. Plants were watered only when soil was dry. In early May, 36 surviving plants were moved outside, covered with a mesh fabric for 4 d, and then randomly transplanted into a field plot. After scape emergence, plants were covered with a mesh cage and flies were introduced for intercrossing. Umbels were combined at harvest, seed were cleaned, and the resulting population was named Cycle#1. Twenty plants of Cycle#1 were grown in each of two greenhouses (blocks) with waxy DH2107 and glossy B9885 in a randomized complete block design. At 8 and 9 weeks after planting, leaves were sampled for wax measurements from each plant as described above.
Bulbs of SG-Syn were produced in field plots, harvested, vernalized as described, and planted in a greenhouse in early February with 13-h fluorescent lighting. As previously described, leaf samples were taken from each plant at 8 and 9 weeks after planting to measure epicuticular waxes. Three groups of two or three plants with the lowest FA·(Hent+H16)−1 ratios and three groups of two to four plants with the highest FA·(Hent+H16)−1 ratios were selected and allowed to flower; umbels were covered with mesh cages and intercrossed using flies to produce six families (low 1, low 2, low 3, high 1, high 2, and high 3) representing one cycle of phenotypic selection for low and high FA·(Hent+H16)−1 ratios. Twelve plants from each of the six selections were grown in the same two greenhouses (blocks) in a randomized complete block design as described above.
Tubes containing waxes dissolved in chloroform were allowed to dry in a fume hood. After the chloroform in a tube had evaporated, 250 µL of anhydrous HPLC chloroform (Sigma-Aldrich) was added, the tube was tightly capped and gently rotated to remove any deposits from sides of the tubes. After all samples had anhydrous chloroform, 300 µL of acetonitrile (Fisher Scientific) and 105 µL of N,O-Bis(trimethylsilyl)trifluoroacetamide (Sigma-Aldrich) were added to each tube. The tubes were tightly capped, derivatized at 80 °C for 30 min, and placed in a shaker at 100 rpm for 5 min. Then, contents were filtered through sterile polyvinylidene fluoride filters with a diameter of 13 mm and pore size of 0.22 μm (Fisher Scientific) into GCMS vials. Samples were injected using an autosampler (PALS RSI 120; Agilent, Santa Clara, CA, USA) into a column (19091S-433UI; Agilent) on a gas chromatography–mass spectrometer (GC model 8890, MS model 5977C; Agilent). All injections were performed using a Hamilton syringe and the split injection method. The initial column oven temperature was 150 °C and the injection temperature was 250 °C. Samples were run on a cycle of 150 °C for 10 min, followed by a ramp-up of the temperature to 300 °C over 10 min and then held at 300 °C for 10 min.
After each GCMS run, a post-run analysis (MassHunter 10.2 SR1 and Qual SW 10.0 software; Agilent) was used to determine peak areas and identities of waxes as described by Khosa et al. (2020). Peak areas were adjusted by dividing by the dry weight of leaf pieces and, therefore, represent peak areas per gram of leaf dry weight. Peak areas from two samples from the same plants were averaged and used for all analyses. RStudio 2022.07.2 + 576 “Spotted Wakerobin” release (R Foundation for Statistical Computing, Vienna, Austria) was used for statistical analyses. Normality of data was assessed using the Shapiro-Wilk normality test, and data that significantly (P < 0.001) deviated from normality were converted using log10 or square-root transformations to normality before analyses. Means and standard deviations presented in the tables were derived from the nontransformed data. Analyses of variance, Pearson correlations, least square means with the Bonferroni correction, and parent–offspring regressions were calculated using RStudio.
The foliage on field-grown plants of B5351 and PIs 255462, 264320, 264327, 321385, 546188, and 546192 were visually compared with the foliage of glossy B9885 and waxy DH2107 (Munaiz et al. 2020), and bulbs were harvested from plants with foliage waxiness intermediate between that of B9885 and DH2107. After vernalization, individual plants from the PIs were crossed with B5351 and/or each other (Table 1) in a complementation test. The visual semi-glossy phenotype of B5351 is controlled by a major QTL on chromosome 5, at which the waxy phenotype shows dominance over semi-glossy foliage (Damon and Havey 2014). If independent loci conditioned the semi-glossy phenotype of B5351 compared to semi-glossy plants from the PIs, then waxy hybrids could be produced. For all crosses, hybrid progenies were semi-glossy, and self-pollination of hybrid plants produced only semi-glossy progenies. This result demonstrates that six sources of the visually semi-glossy phenotype (PIs 255462, 264320, 264327, 321385, 546188, and 546192) possess the same recessive region on chromosome 5 conditioning the semi-glossy phenotype of B5351 (Damon and Havey 2014). SG-Syn was created by intercrossing among 10 F2 progenies from each of eight families from the complementation test (Table 1), followed by a second generation of random mating. All plants of SG-Syn had visually semi-glossy foliage.
Onion accumulates higher amounts of leaf waxes under protected cover compared with that of field-grown plants (Havey 2022); therefore, vernalized bulbs of SG-Syn (n = 72) and waxy DH2107 (n = 6) were grown in a greenhouse. Leaves on all plants of SG-Syn were visually less waxy compared with DH2107. Additionally, GCMS revealed that plants of SG-Syn and DH2107 accumulated low amounts of the alkanes Hept and Non and the ether Eth, as previously reported (Havey 2022; Munaiz et al. 2020). Coefficients of variation for peak areas for these three waxes on plants of DH2107 were high (245%, 58%, and 69%, respectively) (Table 2); therefore, these waxes were excluded from further analyses. Peak areas were larger for the FAs Heptol, Hex, Oct1, and Tri, the alkane Hent, and the ketone H16, and all of these waxes had coefficients of variation less than 25% on the foliage of DH2107 (Table 2). Plants of SG-Syn had significantly less total wax than that of waxy DH2107 (Fig. 1, Table 2), and the correlation among percentages of individual waxes on leaves of SG-Syn and DH2107 was highly significant (P < 0.001) at 0.993, indicating that the semi-glossy phenotype of SG-Syn is attributable to a reduction in total wax as opposed to significant differences for individual waxes. The amounts of FAs Heptol, Hex, Oct1, and Tri were significantly correlated (potentially all products from the acyl reduction pathway), as were amounts of Hent and H16 (products of the decarbonylation pathway) (Table 3). The only nonsignificant correlations were those between the FAs Heptol and Oct1 and between Oct1 and the alkane Hent (Table 3). The total peak area for FAs (Heptol, Hex, Oct1, and Tri) was significantly correlated with amounts of Hent and H16 (0.646; P < 0.001), indicating that the amounts of waxes produced from the two biosynthetic pathways are associated.
Citation: J. Amer. Soc. Hort. Sci. 150, 3; 10.21273/JASHS05479-25
Significant parent–offspring regression coefficients indicate that a trait is heritable. Regressions of parental versus S1 families (n = 44) for amounts of individual waxes, total wax, total FA, Hent+H16, and the ratios of FA·(Hent+H16)−1 and FA·(total wax)−1 were highly significant (P < 0.001), with the exception of Heptol, which was significant at P < 0.05 (Table 4, Fig. 2). Blocks (greenhouses) were occasionally significant (P < 0.05) for some waxes, but no specific greenhouse was consistently different. The amounts of variation explained by the regression models (R2) ranged from lows of 10.8% for Heptol and 35.4% for Hent to ≥50.0% for the other waxes and their ratios (Table 4). These significant parent–offspring regressions indicate that amounts of individual waxes and groups and ratios of specific waxes, and total wax may respond to selection to produce onion populations with different profiles of epicuticular waxes.
Citation: J. Amer. Soc. Hort. Sci. 150, 3; 10.21273/JASHS05479-25
The FA·(Hent+H16)−1 was chosen as the target for selection to alter wax composition on leaves of the SG-Syn population. Plants from four S1 families from SG-Syn with the highest FA·(Hent+H16)−1 ratios (means: 1.10 ± 0.08 vs. 0.79 ± 0.30 for SG-Syn per se) were intercrossed to generate one cycle of S1-family selection (Cycle#1). The FA·(Hent+H16)−1 ratio of Cycle#1 was significantly greater than that of the parental SG-Syn population (Fig. 3, Table 5). The total wax of Cycle#1 was not significantly different from that of SG-Syn. Although not significant, Cycle#1 plants had greater amounts of the FAs and lower amounts of Hent+H16 relative to SG-Syn, resulting in the significantly greater FA·(Hent+H16)−1 ratio (Table 5).
Citation: J. Amer. Soc. Hort. Sci. 150, 3; 10.21273/JASHS05479-25
One cycle of phenotypic selection was completed by crossing among plants of SG-Syn with significantly lower and higher mean FA·(Hent+H16)−1 ratios (Table 6). The coefficient from regression of the parental FA·(Hent+H16)−1 ratios on those of the six selections was highly significant (0.595 ± 0.089; P < 0.001), with an R2 of 25.8%. Plants from all three of the high selections had greater FA·(Hent+H16)−1 ratios than SG-Syn, although only high 1 was significantly (P < 0.05) greater (Fig. 3, Table 5). Selection high 1 had significantly greater total FA relative to SG-Syn and a statistically similar amount of Hent+H16, resulting in the greater FA·(Hent+H16)−1 ratio (Table 5). This indicated that it should be possible to select for greater amounts of waxes from the acyl reduction pathway without necessarily lowering the amounts of waxes from the decarbonylation pathway.
Selection high 2 had significantly greater total wax than SG-Syn because of significantly greater amounts of both FAs and Hent+H16 (Table 5). Because the amounts of both FA and Hent+H16 increased, their ratio was not significantly different from the parental SG-Syn. High 3 was not significantly different from SG-Syn in terms of total wax, total FA, Hent+H16, and FA·(Hent+H16)−1, indicating that there was no significant response to selection.
All three of the low selections had lower FA·(Hent+H16)−1 ratios than SG-Syn, although none was significantly lower (Fig. 3, Table 5). Selection low 1 had the lowest FA·(Hent+H16)−1 ratio attributable to significantly greater amounts of (Hent+H16) relative to SG-Syn. Selections low 2 and low 3 were not statistically different from SG-Syn in terms of total wax, total FA, Hent+H16, and FA·(Hent+H16)−1.
Epicuticular waxes on foliage are important for interactions with insects, diseases, and the environment. The semi-glossy foliage of onion suffers less feeding damage from onion thrips compared with that of plants with waxy foliage (Damon et al. 2014). In a cross between semi-glossy B5351 and waxy plants, two major QTL conditioning the amounts of FAs and H16 were identified (Damon and Havey 2014). However, it was not known if unique wax profiles could be selected using only semi-glossy plants. Amounts of individual waxes, total wax, total FA, Hent+H16, and the ratio of FA·(Hent+H16)−1 on the foliage of semi-glossy onion showed significant parent–offspring regressions, indicating that these traits are heritable (Table 4). Response to phenotypic and S1-family selection schemes were observed (Tables 5 and 6), again supporting heritability of low and high FA·(Hent+H16)−1 ratios. However, more than one cycle of selection may be required to significantly alter wax profiles. Onion plants with unique epicuticular wax profiles should be evaluated to determine feeding damage by onion thrips and, if less damage is found, may allow for the development of thrips-resistant cultivars.
Box plots of peak areas of individual waxes and total wax on the foliage of plants of semi-glossy SG-Syn and the waxy doubled haploid (DH) 2107.
Scatter plots with regression lines and percent coefficients of determination (R2) for peak areas from S1 progenies and mean peak areas of parents for the ratio of total fatty alcohols (FAs) over amounts of Hentriacontane (Hent) and 16-Hentriacontanone (H16) [FA·(Hent+H16)−1] (A) and mean peak areas for total wax (B).
Boxplot of fatty alcohol (FA)·(Hent+H16)−1 for the parental synthetic population (SG-Syn), low and high phenotypic selections, and cycle 1 of S1 family selection. Horizontal lines in boxes are the median. The letter X indicates the mean value. H16 = 16-Hentriacontanone; Hent = Hentriacontane.
Contributor Notes
I gratefully acknowledge the support of the US Department of Agriculture (USDA), Agricultural Research Service, and USDA National Institute of Food and Agriculture grant 2018-51181-28435 from the Specialty Crops Research Initiative, as well as Christy Stewart and Scott Hendricks for technical help.
M.J.H. is the corresponding author. E-mail: mjhavey@wisc.edu.
Box plots of peak areas of individual waxes and total wax on the foliage of plants of semi-glossy SG-Syn and the waxy doubled haploid (DH) 2107.
Scatter plots with regression lines and percent coefficients of determination (R2) for peak areas from S1 progenies and mean peak areas of parents for the ratio of total fatty alcohols (FAs) over amounts of Hentriacontane (Hent) and 16-Hentriacontanone (H16) [FA·(Hent+H16)−1] (A) and mean peak areas for total wax (B).
Boxplot of fatty alcohol (FA)·(Hent+H16)−1 for the parental synthetic population (SG-Syn), low and high phenotypic selections, and cycle 1 of S1 family selection. Horizontal lines in boxes are the median. The letter X indicates the mean value. H16 = 16-Hentriacontanone; Hent = Hentriacontane.
