analysis of dna content of wild and cultured labeo calbasu hamilton, 1822 using nanophotometer

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Analysis of DNA Calbasu (Hamilton

1Research ScholarDepartment of Zoology RGPG College

ABSTRACT Labeo calbasu is an important food fish and commonly known as the “calbasu” or “Black Rohu”. The present study was conducted on DNA extraction and determination of DNA quantity of both male and female of wild and cultured Labeo calbasuNanophotometer. DNA isolation was done and gel electrophoresis was carried out. Extracted DNA was analyzed using nanophotometer (Nanophotometer P330; Implen, Germany) to determine the concentration of DNA and its purity level. Several different methods have been employed in the quantification of nuclear DNA over the past 50 years. The ease of use of this technique not only makes it a feasible option for small volume analysis of DNA but also a practical alternative for spectrophotometric measurement. Significant progress was made to measure micro volume liquid samples (<1µl) in biotechnology & pharmaceutical applications. For the present study both male and female of wild calbasu (from middle Ganga Region) and cultured Labeo calbasu (from culture ponds and/ hatchery) were taken. The value of DNA concentration in female of wild Labeo calbasu was in between 58 ng/µl and of male was in between range of 64ng/µl. The value of DNA concentration in female of cultured Labeo calbasu was 60- 66 ng/µ68- 74 ng/µl. KEY WORDS: DNA Quantity, wild, cultured, Nanophotometer, Labeo calbasu. INTRODUCTION Labeo is a genus of carps in the familyLabeo calbasu locally called Kalibasu or Calbasu or Black rohu, is a freshwater fish species and is the most important carp species next to three Indian Major Carps; Labeo rohita, Catla catla

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6470 | Volume - 3 | Issue – 1 | Nov – Dec 2018

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DNA Content of Wild and Cultured LabeoHamilton, 1822) Using Nanophotometer

Nisha Rana1, Seema Jain2 Research Scholar, 2Associate Professor

Department of Zoology RGPG College, Meerut, Uttar Pradesh, India

is an important food fish and commonly known as the “calbasu” or “Black Rohu”. The present study was conducted on DNA extraction and determination of DNA quantity of both male and

Labeo calbasu using Nanophotometer. DNA isolation was done and gel electrophoresis was carried out. Extracted DNA was analyzed using nanophotometer (Nanophotometer P330; Implen, Germany) to determine the concentration of DNA and its purity level. Several

e been employed in the quantification of nuclear DNA over the past 50 years. The ease of use of this technique not only makes it a feasible option for small volume analysis of DNA but also a practical alternative for spectrophotometric

cant progress was made to measure micro volume liquid samples (<1µl) in biotechnology & pharmaceutical applications. For the present study both male and female of wild Labeo

(from middle Ganga Region) and cultured and/ hatchery)

were taken. The value of DNA concentration in was in between 58 -62

l and of male was in between range of 64- 68 l. The value of DNA concentration in female of

66 ng/µl and of male

DNA Quantity, wild, cultured,

in the family Cyprinidae. locally called Kalibasu or Calbasu or

is a freshwater fish species and is the most important carp species next to three Indian

and Cirrhinus

mrigala. The synonyms used for this species areCyprinus calbas (Hamilton, 1822), calbasu (Day, 1878), Labeo calbasuShebbeare, 1937). It is a popular food fish and also is admired as a sport fish. Recently this fish species has also made its entry in ornamental fish markets of India and abroad. In last few years, the natural populations of this fish species has seriously declined due to fishing and other anthropological reasons. The dorsal profile of Calbasu is more convex than of abdomen.Other identifying features includes; thick and fringed. Two pairs of barbellspair longer than maxillary pair (Rahman, 19pores on snout and eye situated a bit anterior from the half of the head (Bhuiyan, 1964). Caudal peduncle is short an lateral line well marked, scales in size. There are 20 rows of scales before dorsal fin and 22 rows round the caudal 1964). Mouth is moderately wide and Gill openings of Kalibasu is wide avilliform, short and feeble (Bhuiyan, 1964). Colourbody is dark-black but the ventral part light dark. ventral surface of the opercular region is white iris coppery. This fish attains a length of(Talwar and Jhingran, 1991). feeder. It feeds on vegetable matter, crustaceans, insect larvae etc. Labeo calbasuhigher plants 48%, protozoa 12%, crustacean 10%, mollusca 5%, mud and sand 15% (Bhuiyan, 1964). It does inhabit rivers and tributaries. They are also seen in deep pools clear sluggish streams, creeks. It can be reared in ponds and also in tanks (Bhuiyan, 1964 and Rahman, 1989). Of wide distribution in India, it is one of the major Indian carps. It is an important food fish and at several

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Dec 2018

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nd Cultured Labeo Using Nanophotometer

, India

The synonyms used for this species are Hamilton, 1822), Labeo Labeo calbasu (Shaw and

It is a popular food fish and also is admired as a sport fish. Recently this fish species has also made its entry in ornamental fish markets of India and abroad. In last few years, the natural populations of this fish species has seriously declined due to over fishing and other anthropological reasons.

The dorsal profile of Calbasu is more convex than of Other identifying features includes; Lips are

pairs of barbells present, rostal pair longer than maxillary pair (Rahman, 1989). No

ye situated a bit anterior from the 964). Caudal peduncle is

ateral line well marked, scales are moderate in size. There are 20 rows of scales before dorsal fin and 22 rows round the caudal peduncle (Bhuiyan, 1964). Mouth is moderately wide and also inferior. Gill openings of Kalibasu is wide and gill rakers are villiform, short and feeble (Bhuiyan, 1964). Colour of

black but the ventral part light dark. The the opercular region is white iris

coppery. This fish attains a length of nearly 90 cm (Talwar and Jhingran, 1991). Generally it is a bottom feeder. It feeds on vegetable matter, crustaceans,

Labeo calbasu feeds on algae 10%, higher plants 48%, protozoa 12%, crustacean 10%, mollusca 5%, mud and sand 15% (Bhuiyan, 1964). It does inhabit rivers and tributaries. They are also seen in deep pools clear sluggish streams, creeks. It can be

tanks (Bhuiyan, 1964 and

Of wide distribution in India, it is one of the major Indian carps. It is an important food fish and at several

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places is referred to as the "Black Rohu". It is an important game fish in the tanks where it is sand is cultivated along with other species. It thrives better in tanks and lakes than in running waters; can tolerate slightly brackish water also. It does not normally breed in ponds; induced bred by hypophysation. It is essentially a bottom feederattains a length of 90 cm. It can be taken on small flyspoon (Talwar and Jhingran 1991). OBJECTIVE To extract DNA of both male and female of wild and cultured Labeo calbasu. To determine concentration of DNA of both male and female of wild and cultured Labeo calbasuNanophotometer. MATERIALS AND METHODS In the present study, Labeo calbasucollected from different water bodies of Western Uttar Pradesh (Bijnor district, district Hapur and from district Meerut), morphologically identified (with the help of standard literature of Day’s fauna 187578,1889; Jayaram, 1981) and preservedstudies (for DNA isolation and quantification).Approx. 100mg of muscle tissue and fin clip from 2individuals of each species were preserved in 95% ethanol until used and will be kept in molecular analysis. Voucher specimens were preserved in 10% formalin solution. DNA isolation was done by following the method of Ruzzante (1996) with minor modifications. The DNA was diluted to a final concentration of 100ng/µL.electrophoresis was carried out by 1.5-2% agarose gel. The extracted DNA was further analyzed using Nanodrop spectrophotometer (Nanophotometer P330; Implen, Germany) to determine the concentration of DNA and its purity level. Total DNA quantification was carried by nanophotometetrically taking absorbance of 260 and 280nm. A total of of Labeo calbasu of wild and cultured analyzed. RESULTS The present study was conducted on the isolation of DNA and determination of DNA concentration of both male and female of wild and cultured calbasu using Nanophotometer.DNA quantification was done by using nanophotometer and all data was noted (Table 1). The value of DNA concentration in female of wild Labeo calbasu was between 58

International Journal of Trend in Scientific Research and Development (IJTSRD) ISSN: 2456


places is referred to as the "Black Rohu". It is an important game fish in the tanks where it is stocked and is cultivated along with other species. It thrives better in tanks and lakes than in running waters; can tolerate slightly brackish water also. It does not normally breed in ponds; induced bred by hypophysation. It is essentially a bottom feeder. It attains a length of 90 cm. It can be taken on small fly-

To extract DNA of both male and female of wild and

To determine concentration of DNA of both male and Labeo calbasu using

Labeo calbasu fishes were collected from different water bodies of Western Uttar

district, district Hapur and from , morphologically identified (with the

help of standard literature of Day’s fauna 1875-78,1889; Jayaram, 1981) and preserved for molecular

quantification). tissue and fin clip from 2-5

individuals of each species were preserved in 95% ethanol until used and will be kept in -20 °C for molecular analysis. Voucher specimens were

DNA isolation method of Ruzzante et.al.,

modifications. The DNA was diluted to a final concentration of 100ng/µL. Gel

2% agarose gel. The extracted DNA was further analyzed using

hotometer P330; to determine the concentration of

Total DNA quantification was carried by nanophotometetrically taking

A total of 20 samples of wild and cultured fishes were

The present study was conducted on the isolation of DNA and determination of DNA concentration of both male and female of wild and cultured Labeo

Nanophotometer.DNA quantification nanophotometer and all data was

The value of DNA concentration in was between 58-62

ng/µl and of male was between (Fig.3). The value of DNA concentration in female of cultured Labeo calbasu was 60 68- 74 ng/µl (Fig. 4).

Table1. Quantity of DNA in all 20 samples of Labeo calbasu in Nanogram/

Sample No.

Wild Female

Wild Male

1 60 66 2 58 64 3 61 66 4 62 68 5 59 64

Nucleic acids play a major role in growth andevelopment. The amount of de(DNA), the carrier of genetic information, is stable under changing environmental situations and has been used as an indicator of biomass and cell number (Regnault and Luquet 1974). study revealed that the highest content of DNA was seen in female fishes of wild and also of culturedHowever slight change was seen in male fish. The comparative result revealed that the highest content was in cultured fishes, where all optimal conditions were maintained and also in female fishes of culturfishes.

Fig. 1 Quantity of DNA content in Wild in Nanogram/ µl/individual

Fig. 2 Quantity of DNA content in Culturedcalbasu in Nanogram/

Research and Development (IJTSRD) ISSN: 2456-6470

Dec 2018 Page: 907

l and of male was between ranges of 64- 68 ng/µl The value of DNA concentration in female of

was 60 - 66 ng/µl and of male

Quantity of DNA in all 20 samples of in Nanogram/ µl/individual

Cultured Female

Cultured Male

66 68 65 70 60 74 66 72 62 74

Nucleic acids play a major role in growth and development. The amount of deoxyribonucleic acid

), the carrier of genetic information, is stable under changing environmental situations and has been used as an indicator of biomass (Dortch et al. 1983) and cell number (Regnault and Luquet 1974). The study revealed that the highest content of DNA was

en in female fishes of wild and also of cultured. However slight change was seen in male fish. The comparative result revealed that the highest content was in cultured fishes, where all optimal conditions were maintained and also in female fishes of cultured

Fig. 1 Quantity of DNA content in Wild L. calbasu

in Nanogram/ µl/individual

Quantity of DNA content in Cultured L.

in Nanogram/ µl/individual



Fig. 3 Comparative DNA Quantification in nanogram/ µl in all 20 samples of both male and

female L. calbasu DNA measurement can provide valuable tool for monitoring the health and condition of fish.knowledge from such investigations can be used in optimizing and sustaining yield, stock management and conservation of genetic diversity. DISCUSSION Nanodrop spectrophotometry is an extremely powerful technology that allows Quantification of DNA, RNA (A260) and protein (A280) concentrations and sample purity (260/280 ratio) over a large concentration range of 2 - 15,000 ng/L double standards DNA (Pratima et. al,. 2014). DNA concentration in female of wild Labeowas between 58-62 ng/µl and of male was between ranges of 64- 68 ng/µl. The value of DNA concentration in female of cultured Labeowas 60 - 66 ng/µl and of male 68- 74 ng/al., (2012) used nanodrop method quantification from different fishes bastarget. Nanodrop technique was also used by Shi al., (2015) for DNA quantification in the process of molecular characterization of Cynoglossus semilaevis.In the project report submitted by Aderibigbe Adedunni (2014) to the department of aquaculture and fisheries management, University of agriculture, Nigeria determination of DNA concentration of Clarias gariepinus was done by nanodrop. DNA quantification of Male and Female Clarias batrachus, Clarias gariepinus andClarias hybrids was also by Shobhna (2017). She observed the value of DNA concentration in female of C. batrachus between 59 and 61 ng/µl and of male was in between range of 70- 76 ng/µl. The value of DNA concentration in female of C. gariepinus 63 ng/µl and of male 75- 78 ng/µl. Highest DNA concentration was seen in hybrid individuals, in females the concentration was in between 74



Fig. 3 Comparative DNA Quantification in

in all 20 samples of both male and

DNA measurement can provide valuable tool for monitoring the health and condition of fish. The knowledge from such investigations can be used in optimizing and sustaining yield, stock management

Nanodrop spectrophotometry is an extremely powerful technology that allows Quantification of

A (A260) and protein (A280) concentrations and sample purity (260/280 ratio) over

15,000 ng/L double 2014). The value of

Labeo calbasu l and of male was between

l. The value of DNA Labeo calbasu

74 ng/µl. Prado et. method for DNA

quantification from different fishes based on nuclear target. Nanodrop technique was also used by Shi et.

(2015) for DNA quantification in the process of Cynoglossus semilaevis.

In the project report submitted by Aderibigbe aquaculture and

fisheries management, University of agriculture, Nigeria determination of DNA concentration of

was done by nanodrop. DNA Clarias batrachus,

hybrids was also done he value of DNA

C. batrachus was in l and of male was in between

The value of DNA C. gariepinus was 60 and

µl. Highest DNA concentration was seen in hybrid individuals, in females the concentration was in between 74-76 ng/µl

and in case of males value was between 88Similar study on diminution level of RNA/DNA ratio in tissue of Labeo rohita endocrine disrupting compounds was also done by Verma et al., 2016. REFERENCES 1. A. Adedunnijanet, “Isolation and determination of

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analysis of dna content of wild and cultured labeo calbasu hamilton, 1822 using nanophotometer

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