Sex identification in Populus ciliata using morphological characters of leaves Amita Kumari*, Rakesh Kumar Shukla and Prem Kumar Khosla

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1 Sex identification in Populus ciliata using morphological characters of leaves Amita Kumari*, Rakesh Kumar Shukla and Prem Kumar Khosla School of Biological and Environmental Sciences, Shoolini University, Solan (H.P.), India Abstract Corresponding Author Amita Kumari School of Biological and Environmental Sciences, Shoolini University, Solan (H.P.), India amitabot@gmail.com Article History Received on 21 February, 2016; Received in revised form 16 March, 2016; Accepted 30 March, 2016 Morphological variations between the leaves of male and female trees of Populus ciliata were studied in the month of August from Shimla district (H.P.). Different qualitative traits (i.e. shape of the base and tip of the leaf blade, sinus with petiole, pubescence on the lower surface of the leaf blade, leaf margin and colour of the blade) and quantitative traits [i.e. leaf area (LA), lamina length/width (L/W) percent, petiole length/ nerve length (P/N) percent, perimeter, aspect ratio (AR) and shape factor (SF)] were recorded from the thirty leaves of four male and four female trees. Results showed that leaves of male and female trees of P. ciliata were differentiated on the basis of three qualitative characters (shape of base of leaf blade, sinus with petiole and leaf margin). Quantitative parameters showed non-significant results except the shape factor which was found to be statistically significant, therefore able to distinguish the two sexes of P. ciliata. Keywords : Morphological variations, quantitative, qualitative and Populus ciliata Introduction Populus ciliata Wall. ex Royle is a dioecious tree species in which male and female flowers are borne on different trees. A few studies have recommended the use of male cultivars over the female ones as males are more adapted to xeric sites whereas females prefer to grow on fertile sites (Khosla et al., 1979). The female clones have more biomass than the male clones due to the production of more lateral branches (Tschaplinski et al., 1994). In the vicinity of habitations, propagation of male plants are recommended because silky and light seeds become an aesthetic and health hazard. The gender on this plant can only be identified at the flowering stage, which usually take five to seven years and therefore create problems in plant breeding as well as its clonal propagation. Morphologically male and female trees are indistinguishable from each other on the basis of vegetative buds and the growth rate. No information is available on a morphological basis of the gender determination on poplar plants and efforts has only been made in the genetic or molecular level. Traditionally, morphological characters have been used for cultivar identification and rarely they have been able to identify the male and female trees in the field except at the flowering stage. Materials and Methods To solve the present problem, thirty leaves from four male and four female trees of P. ciliata were collected from the district Shimla (near Sankat Mochan Temple), H.P. The sampling site is located on 31 06' N/ 77 13' E, northern India has average elevation 1975 m above sea level. Male and female trees were Int. J. Adv. Lif. Sci., Available online on at www. ijals.com Page 174

2 identified and marked during the flowering season (March - April) prior to sample collection. The average of thirty observations was taken using statistical tools ( t-test; Prism software). The following different morphological characters were studied. Qualitative characters (Vasudha, 2000): General shape of base of leaf blade; General shape of tip of leaf blade; Shape of sinus with petiole; Pubescence on the lower surface of leaf blade; Serration of the edge of the leaf lamina and colour of leaf blade. Quantitative characters (Anonymous, 2012; Rabha et al., 2013) : Leaf area (LA) in cm 2 ; Perimeter of leaf; Average value in percent for the ratio of lamina length and width (L/W %); Average value in percentage for the ratio of petiole to medial nerve (P/N %); Aspect ratio (AR) and Shape factor (SF). Results and Discussion Qualitative study In the present study efforts have been made to identify the male and female trees of P. ciliata through qualitative traits on the basis of guidelines laid out by the International Poplar Commission (IPC). The results so far obtained are discussed here under: The table -1 summarises the morphological shape of the base and tip of the leaf blade, sinus with petiole, pubescence on the lower surface of the leaf blade, leaf margin and colour of the blade of male and female P. ciliata trees. The differences between male and female trees were seen in the shape of the leaf blade being deltoids-cordate in females and deltoids-ovate in males. Similarly, the shape of the sinus with petiole was deep in females while shallow in males and undulation of the leaf blade margin was sparsely serrate in females and densely serrate in males (Fig.-1). These phenotypic traits were found to be identifying characteristics of female and male trees of P. ciliata. Such type of variation in many traits were also observed by Jokela et al. (1976) and Mohanty and Khurana (2000). These results concluded that morphological variation (especially qualitative traits, i.e. shape of leaf lamina, serration of the edges of leaf blade, shape of the sinus with petiole, etc.) exists between the male and female trees of P. ciliata and could be used to differentiate the gender of the trees at vegetative stage. Although, contradictory results have been observed in Myristica fragrans (Nayar et al., 1977 and Zachariah et al., 1986,) and Telfairia occidentalis (Ndukwu et al., 2005) where morphological characteristics of leaves were unable to differentiate gender of the plant. Quantitative study Different quantitative characters studied were Leaf Area (LA), Lamina length/ Width (L/W %), Petiole length / Nerve length (P/N %), Perimeter, Aspect Ratio (AR) and Shape factor (SF), shown in table - 2. Male in general excelled the females in all characters enumerated in this study except the L/W % where in males, ( ) it was marginally lower than the females ( ). The values of average LA, P/N %, perimeter, AR and SF in male genotypes were , , , and , whereas in female genotypes were , , , and respectively (Fig.- 2). The variations in these characters were due to the result of existing variation in the leaf blade length, blade width, nerve length and petiole length, which ultimately contributed to the wide range of variation in leaf area observed for male trees of P.ciliata. Although, results of leaf area, P/N%, perimeter of the leaf and aspect ratio showed small variations between male and female trees leaf of P. ciliata but statistically (t -test) these variations were not significant. Only the result obtained within a shape factor was found statistically significant (t - test; P <0.0001) (Table - 3). Robinson et al. (2014) also Int. J. Adv. Lif. Sci., Available online on at www. ijals.com Page 175

3 Table - 1. Qualitative characters of male and female tree leaves of P.ciliata Sr. No. Characters of leaf Females Males 1. General shape of base of leaf blade Deltoido- cordate Deltoido-ovate 2. Shape of sinus with petiole Deep Shallow 3. General shape of tip of leaf blade Acute Acute 4. Pubescence on lower surface of leaf blade 5. Undulation of the edge of leaf blade (leaf margin) On whole surface Sparsely serrate 6. Colour of leaf blade Green on upper surface and whitish on lower surface Table - 2. Quantitative characters of male and female tree leaves of P. ciliata tree Sr. No. Leaf Characters On whole surface Densely serrate Green on upper surface and whitish on lower surface P. ciliata Mean (SD) Male trees Female trees 1. Leaf Area (LA) ( ) ( ) 2. Lamina length/ Width % (L/W %) ( ) ( ) 3. Petiole length/ Nerve length % (P/N %) ( ) ( ) 4. Perimeter ( ) ( ) 5. Aspect Ratio (AR) ( ) ( ) 6. Shape factor (S/F) ( ) ( ) Table - 3. t tests between leaf area, leaf length/width % and leaf petiole/ middle nerve percent, perimeter of leaf, aspect ratio and shape factor of male and female trees of P. ciliata. t - test Mean Difference t value P value Significant Leaf area (cm 2 ) Male vs Females ± ns Leaf length/ width % Males vs females ± ns Leaf petiole/middle nerve length % Males vs females ± ns Perimeter of leaf Males vs Females ± ns Aspect ratio Males vs Females ± ns Shape factor Males vs Females ± < *** *** - Significant (P<0.0001), ns- Non-significant (P>0.05) Int. J. Adv. Lif. Sci., Available online on at www. ijals.com Page 176

4 Fig Leaf of female (a) and male (b) tree of P. ciliata from abaxial and adaxial surface reported non-significant difference between the size of male and female tree leaves of P. tremula. All these studied quantitative characters (LA, Lamina length, Lamina width, petiole length, etc.) during the study have been previously used for genotype determination in Chestnut (Serdar and Kurt, 2011). Wheelwright et al. (2012) observed non-significant result with the higher leaf area in Ocotea tenera females. Li et al. (2007) observed higher specific leaf area in females of Hippophae rhamnoides than males. Durako and Moffler (1985) compared the leaf width between the sexes of Thalassia testudinum and observed narrow leaves of female than males. The leaf area plays an important role in determining the growth rate of the plant through net photosynthetic rate. Wang and Curtis (20 10) observed higher photosynthetic rate in male saplings of Populus tremuloides than females throughout the growing season. The growth rate of leaves in female trees of P. ciliata was observed minimum because female expends more energy in its reproduction and seed maturation, therefore having less growth rate and minimum leaf area than males as observed during the study. Such type of correlation between growth rate and leaf surface was suggested by Dickmann (1971) and Mohanty and Khurana (2000) in poplar trees. Similar types of relationship between leaf area of blade Fig-2. Variation between the mean average value of LA (A.), L/W % (B.), P/N % (C.), AR (D.) leaf perimeter (E.) and SF (F.) of male and female trees of P. ciliata. and length-breadth was also observed by Narkhede (1995) in poplar plants in consonance with the present study. The other reason of the lower leaf area in females could be that the trees selected for present investigation were offering a stressful environment resulting in a negative effect on the females than males (Li et al., 2007 and Xu et al., 2008). The growth characteristics in P. ciliata are controlled by both genetic and environmental factors (Khurana et al., 1992 and Khosla et al., 1979) which is proved by the present quantitative study. On the other hand qualitative characters are observed to be fixed, controlled by only the genotype of the trees and can be further used for sex identification purposes in the field at the mature stage of their growth. This is the small effort to identify the morphological variations between male and females of P. ciliata but the study require the use of a large number of morphological characters on number of male and female trees along the latitudinal and altitudinal gradient. Int. J. Adv. Lif. Sci., Available online on at www. ijals.com Page 177

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6 Wheel wright, N.T., Sinclair, J.P., Hochwender, C. and Janzen, F.J Leaf size in three generations of a dioecious tropical tree, Ocotea tenera (Lauraceae): sexual dimorphism and changes with age. Am. J. Bot., 99 : Xu, X., Yang, X., Zhang, S., Korpelainen, H. and Li, C Sex-specific responses of Populus cathayana to drought and elevated temperatures. Plant Cell Environ., 31: Acknowledgement Authors are thankful to Dr. Poonam Shirkot, Professor, Dr.Y.S. Parmar University of Horticulture and Forestry, Solan (H.P.), India for her unreserved help during research work. Zachariah, J.A., Krishnamurthy, G.B. and Ravindran, P.N Steroid degradation compound associated with sex expression in nutmeg (Myristica fragrans Houtt.). Proc. Indian Nat. Sci. Acad., 52 : Corresponding Author : Amita Kumari, School of Biological and Environmental Sciences, Shoolini University, Solan (H.P.), India. amitabot@gmail.com 2016, IJALS. All Rights Reserved. Int. J. Adv. Lif. Sci., Available online on at www. ijals.com Page 179