RAPD-PCR based genomic characterization of two populations of Culex quinquefasciatus (Diptera : Culicidae)
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Abstract
The present paper deals with the RAPD-PCR based genomic characterization of Culex quinquefasciatus Say which is a major vector of filariasis in several parts of the Indian subcontinent. One population of the test organism used in the study was procured from Goa (pop.A) while the other (pop.B) was collected from a village Nadasahib (20 kms from Chandigarh). The RAPD-PCR amplification of whole body homogenate of freshly hatched individual specimens was carried out by using three random primers: primer I- 5’- GTCCCGACGA – 3’; primer II- 5’
– TGATCCCTGG – 3’ and primer III- 5’- GTGACGTAGG – 3’. Primer I produced 5 distinct bands from the DNA of pop. A, whose base composition ranged from 200-1000 bp. Likewise, 7 bands ranging from 130-750 bp and 4 bands ranging from 270-950 bp were generated with primers II and III respectively. In case of pop.B, a total of 8 bands ranging from 200-1000 bp were generated with primer I. Similarly, a total of 6 bands ranging from 250-900 bp with primer II and 4 bands ranging from 180-950 bp with primer III were produced. Based on the band
sharing coefficient and the application of Nearest Neighbour Joining (NJ) analysis it was found that primer Iwas more suitable for detecting genomic differences at the species and generic levels while primer II was ideal for detecting variations in the number of bp in RAPD generated bands among different populations of Cx. quinquefasciatus.
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Keywords
PCR, Cx. quinquefasciatus, Genomics
References
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Beebe, N.W., Cooper, R.D., Foley, D.H. and Ellis, J.H. (2000). Populations of the south-west Pacific malaria vector Anopheles farauti s.s. revealed by ribosomal DNA transcribed spacer polymorphisms. Heredity, 84: 244- 253.
Bhargavi, R., Vishwakarma, S. and Murty, U.S. (2005). A secondary structural common core in the ribosomal ITS2 (internal transcribed spacer) of Culex species from diverse geographical locations. Bioinformation, 1(2): 52-55.
Chaudhry, S. (1999). A review of polytene chromosome dynamics in genus Anopheles (Diptera: Culicidae). In: R.C. Sobti and J.S. Yadav (Eds.) Some aspects on the insight of insect biology (pp 205-236), New Delhi: Tausco Book Distributors.
Collins, F.H. and Paskewitz, S.M. (1996). A review of the use of ribosomal DNA (rDNA) to differentiate among cryptic Anopheles species. Insect Molecular Biology, 5(1): 1-9.
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Severini, C., Silverstrini, F., Mancini, P., La Rosa, G. and Marinucci, M. (1996). Sequence and secondary structure of the rDNA second internal transcribed spacer in the sibling species Culex pipiens L. and Cx. quiquefasciatus Say (Diptera: Culicidae). Insect Molecular Biology, 5(3): 181-186.
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Stevens, N.M. (1910). The chromosomes in the germ cells of Culex. Journal of Experimental Zoology, 8: 207-217.
UC Davis News Information (2002). Mosquito Ecology Needs Study of Genetically Modified Mosquitoes are to Curb Disease. pp 1-2. Retrieved May 22, 2009. http://www.news.ucdavis.edu/search/news_detail.
Urdaneta, L., Herrera, F., Pernalete, M., Zoghbi, N., Rubio- Palis, Y., Barrios, José Rivero, R., Comach, G., Jiménez, M. and Salcedo, M. (2005). Detection of dengue viruses in field-caught Aedes aegypti (Diptera: Culicidae) in Maracay, Aragua state, Venezuela by type-specific polymerase chain reaction. Infection Genetics and Evolution, 5(2): 177-184.
Wilkerson, R.C., Parsons, T.J., Albright, D.G., Klein, T.A. and Braun, M.J. (1993). Random amplified polymorphic DNA (RAPD) markers readily distinguish cryptic mosquito species (Diptera: Culicidae: Anopheles). Insect Molecular Biology, (4): 205-211.
Wilkerson, R.C., Parsons, T.J., Klein, T.A., Gaffigan, T.V., Bergo, E. and Consolim, J. (1995). Diagnosis by random amplified polymorphic DNA polymerase chain reaction of four cryptic species related to Anopheles (Nyssorhynchus) albitarsis (Diptera: Culicidae) from Paraguay, Argentina, and Brazil. Journal of Medical Entomology , 32(5): 697-704.
WHO/UNDP/World Bank, (2003). Special Programme for Research and Training in Tropical Diseases (TDR). Molecular Entomology. 1-5.
Ballinger-Crabtree, M.E., Black, W.C. 4th and Miller, B.R. (1992). Use of genetic polymorphisms detected by the random-amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) for differentiation and identification of Aedes aegypti subspecies and populations. The American Journal of Tropical Medicine and Hygiene, 47: 893-901.
Beebe, N.W. and Saul, A. (1995). Discrimination of all the members of the Anopheles punctulatus complex by polymerase chain reaction – restriction fragment length
polymorphism analysis. The American Journal of Tropical Medicine and Hygiene, 53: 478-481.
Beebe, N.W., Cooper, R.D., Foley, D.H. and Ellis, J.H. (2000). Populations of the south-west Pacific malaria vector Anopheles farauti s.s. revealed by ribosomal DNA transcribed spacer polymorphisms. Heredity, 84: 244- 253.
Bhargavi, R., Vishwakarma, S. and Murty, U.S. (2005). A secondary structural common core in the ribosomal ITS2 (internal transcribed spacer) of Culex species from diverse geographical locations. Bioinformation, 1(2): 52-55.
Chaudhry, S. (1999). A review of polytene chromosome dynamics in genus Anopheles (Diptera: Culicidae). In: R.C. Sobti and J.S. Yadav (Eds.) Some aspects on the insight of insect biology (pp 205-236), New Delhi: Tausco Book Distributors.
Collins, F.H. and Paskewitz, S.M. (1996). A review of the use of ribosomal DNA (rDNA) to differentiate among cryptic Anopheles species. Insect Molecular Biology, 5(1): 1-9.
Cornel, A.J., Porter, C.H. and Collins, F.H. (1996). Polymerase chain reaction species diagnostic assay for Anopheles quadrimacualtus cryptic species (Diptera: Culicidae) based on ribosomal DNA ITS2 sequences. Journal of Medical Entomology, 33(1): 109- 116.
Crabtree, M.B., Savage, H.M. and Miller, B.R. (1995). Development of a species diagnostic PCR assay for the identification of Culex vectors of St. Louis encephalitis virus based on interspecies sequence variation in ribosomal DNA spacers. The American Journal of Tropical Medicine and Hygiene, 53(1): 105-109.
Hill, S.M. and Crampton, J.M. (1994). DNA based methods for the identification of insect vectors. Annals of Tropical Medicine and Hygiene, 88: 227-250.
Holt, R.A., Subramanian, G.M., Halpern, A., Sutton, G.G., Charlab, R., Nusskern, D.R. (2002). The genome sequence of the malaria mosquito Anopheles gambiae. Science, 298(55951): 129-149.
Lanzaro, G.C., Nuzhdin, S. and Tripet, F. (2009). Tools for monitoring the genetic structure and stability of mosquito populations. pp 157-164. Retrieved April 20, 2009 from: gelanzaro@ucdavis.edu.
Li, C. and Wilkerson, R.C. (2005). Identification of Anopheles (Nyssorhynchus) albitarsis complex species (Diptera: Culicidae) using rDNA ITS2-based polymerase chain reaction primers. Memorias do Instituto Oswaldo Cruz, 100(5): 495-500.
Manonmani, A.M., Townson, H., Adeniran, T., Jambulingam, P., Sahu, S. and Vijayakumar, T. (2001). rDNA-ITS2 polymerase chain reaction assay for the sibling species of Anopheles fluviatilis. Acta Tropica, 78(1): 3-9.
Proft, J., Maier, W.A. and Kampen, H. (1999). Identification of six sibling species of the Anopheles maculipennis complex (Diptera: Culicidae) by a polymerase chain reaction assay. Parasitology Research, 85(10): 837-843.
Rutledge, C.R., Wesson, D.M. and Meek, C.L. (1999). Polymerase chain reaction assay to identify all immature stages of two species of the Anopheles quadrimaculatus sibling species complex (Diptera: Culicidae). Journal of the American Mosquito Control Association, 15(4): 573-575.
Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989). Molecular cloning: a laboratory manual, 2nd ed. New York: Cold Spring Harbor.
Severini, C., Silverstrini, F., Mancini, P., La Rosa, G. and Marinucci, M. (1996). Sequence and secondary structure of the rDNA second internal transcribed spacer in the sibling species Culex pipiens L. and Cx. quiquefasciatus Say (Diptera: Culicidae). Insect Molecular Biology, 5(3): 181-186.
Singh, O.P., Chandra, D., Nanda, N., Raghavendra, K., Sunil, S., Sharma, S.K., Dua, V.K. and Subbarao, S.K. (2004). Differentiation of members of the Anopheles fluviatilis species complex by an allele-specific polymerase chain reaction based on 28S ribosomal DNA sequences. The American Journal of Tropical Medicine and Hygiene, 70(1): 27-32.
Stevens, N.M. (1910). The chromosomes in the germ cells of Culex. Journal of Experimental Zoology, 8: 207-217.
UC Davis News Information (2002). Mosquito Ecology Needs Study of Genetically Modified Mosquitoes are to Curb Disease. pp 1-2. Retrieved May 22, 2009. http://www.news.ucdavis.edu/search/news_detail.
Urdaneta, L., Herrera, F., Pernalete, M., Zoghbi, N., Rubio- Palis, Y., Barrios, José Rivero, R., Comach, G., Jiménez, M. and Salcedo, M. (2005). Detection of dengue viruses in field-caught Aedes aegypti (Diptera: Culicidae) in Maracay, Aragua state, Venezuela by type-specific polymerase chain reaction. Infection Genetics and Evolution, 5(2): 177-184.
Wilkerson, R.C., Parsons, T.J., Albright, D.G., Klein, T.A. and Braun, M.J. (1993). Random amplified polymorphic DNA (RAPD) markers readily distinguish cryptic mosquito species (Diptera: Culicidae: Anopheles). Insect Molecular Biology, (4): 205-211.
Wilkerson, R.C., Parsons, T.J., Klein, T.A., Gaffigan, T.V., Bergo, E. and Consolim, J. (1995). Diagnosis by random amplified polymorphic DNA polymerase chain reaction of four cryptic species related to Anopheles (Nyssorhynchus) albitarsis (Diptera: Culicidae) from Paraguay, Argentina, and Brazil. Journal of Medical Entomology , 32(5): 697-704.
WHO/UNDP/World Bank, (2003). Special Programme for Research and Training in Tropical Diseases (TDR). Molecular Entomology. 1-5.
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How to Cite
RAPD-PCR based genomic characterization of two populations of Culex quinquefasciatus (Diptera : Culicidae). (2009). Journal of Applied and Natural Science, 1(2), 269-274. https://doi.org/10.31018/jans.v1i2.69
