The Application of UTY and SRY Molecular Markers for Determination of Unknown Sex Samples in Bali Cattle

An investigation involving large number of animals is often resulting incomplete or in accurate information such as animal parentage, or misidentify on sex due to unlabeled sex samples. A PCR method by applying Y chromosome markers (UTY and SRY) facilitates in determination of unknown sex problem. This study was intended to determine sex from unlabelled sex of blood samples by applying PCR method using a pooled-DNA template. Twenty five of unknown sex blood samples from Nusa Penida, Bali were used in this study. The samples were plotted into 5 pooled-DNA whith each pool DNA consisted of 5 individuals DNA. Two pairs of sex primers, UTY (58oC) and SRY (60oC) with 35 cycles were applied to amplify the samples. The result showed there was only one pooled-DNA (P4) amplified by UTY (484bp). Whereas re-PCR of the positive pooled-DNA (P4) using SRY primer, only one out of 25 samples determined as male Bali cattle (325bp). This finding suggests that UTY and SRY primers are suitable for sex determination and the pooled-DNA could be used as an efficient PCR method both in consumables and PCR process for sex determination. Keywords: Determination, sex, unknown sample, pooled DNA, Bali cattle.

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PENENTUAN JENIS KELAMIN BURUNG KENARI (Serinus canaria) BERDASARKAN GEN Chromodomain Helicase DNA-Binding 1 (CHD1)

Jurnal Bioteknologi & Biosains Indonesia (JBBI) ◽

10.29122/jbbi.v7i1.3178 ◽

2020 ◽

Vol 7 (1) ◽

Author(s):

Afif Muhammad Akhrom ◽

Indarjulianto Soedarmanto ◽

Yanuartono Yanuartono ◽

Trini Susmiati ◽

Alfarisa Nururrozi ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Sex Determination ◽

Mature Female ◽

Mature Male ◽

Chain Reaction ◽

Blood Samples ◽

Pcr Method ◽

Polymerase Chain ◽

Female Canaries

Phenotype determination of sex in young canaries is very low in accuracy. This study aimed to develop a genotypic sexing method in canaries. This study used 12 canaries consisting of 3 mature males, 3 mature females and 6 one-month-old canaries. Phenotypic sexing by cloacal observation was done on all birds, continued by genotypic sexing to identification CHD1 gene using polymerase chain reaction (PCR). The PCR used blood samples for mature canaries, and feather for mature and one-month-old canaries. The results of phenotypic observations showed that all mature male canaries had prominent and pointed cloaca forms, all mature females had flat and wide, whereas all one-month-old birds had a flat cloaca. The result of PCR showed a single band (500 bp) for mature male and double bands (500 bp and 300 bp) for mature female canaries. The PCR results of one-month-old canaries showed that there were one male and five females. Based on this study, it was concluded that genotypic sexing using the PCR method is effective in the sex determination of canaries.Keywords: canary, CHD1, genotype, PCR, sexingÂ ABSTRAKPenentuan jenis kelamin burung kenari muda secara fenotip akurasinya sangat rendah. Penelitian ini bertujuan untuk menentukan jenis kelamin burung kenari secara genotip. Penelitian ini menggunakan 12 ekor burung kenari, terdiri dari 6 ekor dewasa (3 jantan, 3 betina) serta 6 ekor umur 1 bulan. Semua burung ditentukan jenis kelaminnya dengan mengamati kloaka dan identifikasi gen CHD1 menggunakan teknik polymerase chain reaction (PCR). Sampel DNA berasal dari darah dan bulu untuk burung dewasa serta bulu untuk burung umur 1 bulan. Pengamatan fenotip menunjukkan bahwa burung kenari dewasa jantan mempunyai bentuk kloaka menonjol dan runcing, dewasa betina berbentuk datar dan lebar, sedangkan semua burung umur 1 bulan mempunya bentuk kloaka datar. Hasil identifikasi gen CHD1 diperoleh adanya 1 pita gen sekitar 500 bp dari sampel darah dan bulu semua burung kenari dewasa jantan, dan 2 pita gen sekitar 500 bp dan 300 bp dari sampel semua burung kenari betina dewasa. Hasil PCR pada sampel burung umur 1 bulan menunjukkan bahwa 1 ekor jantan dan 5 ekor betina. Berdasarkan penelitian ini dapat disimpulkan bahwa penentuan jenis kelamin secara genotip menggunakan gen CHD1 dapat dilakukan pada burung kenari.

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Assessment of seed quality and sex determination in hop seedlings using molecular markers

Ciência e Natura ◽

10.5902/2179460x43135 ◽

2020 ◽

Vol 42 ◽

pp. e45

Author(s):

Marília Pereira Machado ◽

Andreza Cerioni Belniaki ◽

André Felipe Bernert ◽

Erik Nunes Gomes ◽

João Carlos Bespalhok Filho ◽

...

Keyword(s):

Molecular Markers ◽

Sex Determination ◽

Seed Quality ◽

Germination Percentage ◽

Brewing Industry ◽

Breeding Programs ◽

Normal Seedling ◽

Physiological Quality ◽

Male Specific

Brazil is the world's third largest beer consumer and currently imports all of its hops for the brewing industry. Such a fact justifies the selection of hop genotypes adapted for cultivation locally, which requires high quality seeds and efficient sex determination of the seedlings. The objectives of this study were to develop a methodology to assess hop seed quality and to efficiently determine hop seedling sex through the use of male-specific molecular markers. Freshly harvested hop seeds were germinated with and without pre-chilling (3-5 ° C) for 3, 6 and 12 weeks and then germinated at 20 or 25 ° C in the presence or absence of light, evaluating germination percentage and germination speed index. F1 progenies were obtained from after seed germination in a greenhouse and seedlings sex was determined using male-specific molecular markers. The best conditions for physiological quality assessment of hop seeds used in the present study were pre-chilling for 12 weeks, followed by germination at 25 ° C, and normal seedling counts at 7 and 15 days. The progeny submitted to molecular marker sexing was composed of 61.3% female plants. The established methodologies presented here can be considered efficient and may contribute to expedite hops breeding programs.

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Comparative Evaluation Of Two Techniques Of Sex Determination In Lovebirds (Agapornis Spp.)

Bulletin of University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca Veterinary Medicine ◽

10.15835/buasvmcn-vm:2020.0030 ◽

2020 ◽

Vol 77 (2) ◽

pp. 106

Author(s):

Maria Carmen TURCU ◽

Lucia Victoria BEL ◽

Tommaso COLLARILE ◽

Dana Liana PUSTA

Keyword(s):

Sex Determination ◽

Comparative Evaluation ◽

Surgical Instruments ◽

Dna Testing ◽

Blood Samples ◽

Non Invasive ◽

Dna Sampling ◽

Invasive Method ◽

Pcr Method ◽

Materials Used

Most lovebirds (Agapornis spp.) have no distinct sexual dimorphic traits. The objective of the paper was to compare the results of two sexing methods, surgical sexing by celioscopy and DNA sexing by PCR from blood samples, in order to evaluate their effectiveness. Materials used to carry out the studies were a 2.7 mm telescope and endoscopy unit, surgical instruments and DNA sampling kits provided by Exomed.cz laboratory. Forty-two lovebirds were included in this study. The endoscopic sexing procedure was performed by the method described by Divers. Blood sampling for DNA testing by PCR method was performed from the metatarsal vein. The results were different in the case of one lovebird, endoscopic sexing revealed it as male, compared to DNA testing, where it was identified as female. DNA sexing is a non-invasive method that might be more accurate than celioscopy in this species, and bird owners have easier access to it.

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SEX DETERMINATION OF KIWIFRUIT SEEDLINGS WITH MOLECULAR MARKERS

Acta Horticulturae ◽

10.17660/actahortic.2014.1048.24 ◽

2014 ◽

pp. 197-203

Author(s):

A. Atak ◽

B. Aydin ◽

K. Abdurrahim Kahraman

Keyword(s):

Molecular Markers ◽

Sex Determination

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Exon 9 LEPR Gene SNP Polymorphism of Hybrid Chickens F2Kambro Crossbreeds of ♀ F1Kambro with ♂ F1Kambro

10.1101/2021.02.13.431072 ◽

2021 ◽

Author(s):

I Wayan Swarautama Mahardhika ◽

Budi Setiadi Daryono

Keyword(s):

Nucleotide Polymorphisms ◽

Arms Pcr ◽

Specific Protocol ◽

Correlation Degree ◽

Transversion Mutation ◽

Gene Correlation ◽

Exon 9 ◽

Pcr Method ◽

Dna Template

AbstractThe implementation of the T-ARMS PCR method in the detection of single nucleotide polymorphisms (SNPs) in the LEPR gene in chicken DNA samples has never been conducted. This research aims to design a specific protocol for exon 9 LEPR gene SNPs detection and detect LEPR gene expression or LEPR SNPs in Pelung chicken samples, F1Pelung, Layer, Broiler Cobb 500, F1Kambro chicken and F2Kambro chicken using the T-ARMS PCR method. Determination of LEPR gene correlation degree on Body Weight (BT) and Egg Productivity (PT) in F1 Kambro population and F2Kambro. Qualitative phenotype parameters showed six groups of segregated phenotypes compared to F1Kambro chicken. Growth of F2Kambro chicken weight reached 753.36 ± 155.31 grams in 8 weeks was not significant for F1Kambro chicken due to inbreeding depression (Fx = 25%, IR = 4.925%) and transversion of A LEPR allele mutations. Specific protocol detection of exon 9 LEPR gene SNPs using the T-ARMS PCR method can detect C127A LEPR mutations with IP: OP ratio 10:1 pmol / µM, chicken DNA template concentration of 100 ng / µL with annealing temperature of 55.7° C / 30s. The transversion mutation of C127A of LEPR exon 9 SNP were detected in DNA samples of F1Kambro hens (80%), F2Kambro roosters (20%), Broiler Cobb 500 hens (75%). The mutations were not detected in Layer, Pelung Blirik Hitam chicken and F1Pelung populations.

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Molecular sexing in the formation of pairs of blue-and-yellow macaw (Ara ararauna) in reintroduction programs

Research Society and Development ◽

10.33448/rsd-v10i10.19330 ◽

2021 ◽

Vol 10 (10) ◽

pp. 570101019330

Author(s):

Douglas Campos Pereira ◽

Bernardo Mirabal ◽

Adriele Maria Machado de França ◽

Edma Santos de Antonio ◽

Ricardo Evangelista Fraga ◽

...

Keyword(s):

Molecular Markers ◽

Genomic Dna ◽

Low Complexity ◽

Wild Animal ◽

Molecular Sexing ◽

Agarose Gel ◽

Blood Samples ◽

Allele Specific ◽

Males And Females

Blue and yellow macaw is a species which does not show sexual dimorphism and is threatened by animal traffic. The identification of heterosexual pairs is important for reintroduction programs. The aim of this work was select parameters for sexing and use them to determine the frequency of heterosexual pairs in a population of blue-and-yellow macaws allocated in a Wild Animal Screening Center. Blood samples from 23 macaws were collected and genomic DNA extracted by Tris/SDS washes. Allele-specific molecular markers for sexing were amplified by PCR, and identified on 2% agarose gel. Three pairs of primers were tested: Pair 1 (P2/P8), Pair 2 (1237L/1272H) and Pair 3 (2550F/2718R). For the determination of animal pairs, all individuals had their social behavioral acts observed. The results showed that the low complexity DNA extraction protocol used was adequate. Pairs 2 and 3 of primers were effective for sexing and the Pair 3 was the most efficient. The study also showed that in the sample studied, the composition of males and females was similar (0.4 males n=10 and 0.6 females n=13); 70% (n=16) of the individuals formed pairs and 75% (n=12) of the pairs were heterosexual and the others male-male or female-female pairs. These results were used in the management of the animals in the reintroduction program.

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Development of a method for the determination of the JAK2 gene mRNA in venous blood and assessment of its diagnostic value in oncohematology

Russian Clinical Laboratory Diagnostics ◽

10.51620/0869-2084-2021-66-6-379-384 ◽

2021 ◽

Vol 66 (6) ◽

pp. 379-384

Author(s):

M. A. Stolyar ◽

A. S. Gorbenko ◽

I. A. Olkhovskiy ◽

V. I. Bakhtina ◽

M. A. Mikhalev ◽

...

Keyword(s):

Multiple Myeloma ◽

Venous Blood ◽

Diagnostic Value ◽

Janus Kinase ◽

Rt Pcr ◽

Blood Samples ◽

Dnase Treatment ◽

Pcr Method ◽

Gene Mrna

Overactive JAK pathway signaling is a hallmark of immune diseases and critically affects on inflammation and coagulation. A number of mutations in the JAK2 gene act as driving forces of myeloproliferative neoplasms (MPN), the pathogenesis of certain variants of acute leukemia, a number of solid malignancies and cardiovascular diseases. Assays for quantifying JAK2 mRNA in circulating blood cells can be used as a marker associated with the activity of this enzyme. Development of an original method for detecting JAK2 mRNA in venous blood and assessment of the possible diagnostic value in chronic oncohematological diseases. The development of an RT-PCR method for determining the expression of the JAK2 gene mRNA in venous blood samples was carried out in accordance with the MIQE requirements. Primers and TaqMan probes were designed using the Primer3 program, taking into account the possibility of excluding subsequent DNase treatment. The stability of the investigated mRNA was assessed in vacutainers with different anticoagulants and depending on the storage time of the samples. The study of the expression of JAK2 mRNA in blood leukocytes of 41 patients with B-CLL, 16 patients with CML, 12 patients with multiple myeloma and 39 donors using the developed “real-time” PCR method. The study revealed a decrease in the level of JAK2 mRNA in venous blood samples in patients with primary CLL, but not with CML or with multiple myeloma. The level of the marker in the majority of patients with CLL after the start of therapy returned to the range typical for healthy people. It has been shown that the values of the relative expression of JAK2 mRNA are most stable in the range of 2 - 7 hours after taking blood in a vacutainer with EDTA. An original RT-PCR method was developed for the quantitative determination of JAK2 mRNA in venous blood samples, which meets the requirements of the MIQE system. Determination of JAK2 mRNA can be useful for clarifying the pathogenesis features of certain diseases involving impaired Janus kinase activity and can become a promising marker for prognosis and assessment of the effectiveness of therapy.

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DNA sexing for gender determination of Changeable Hawk-Eagle (Nisaetus cirrhatus, Gmelin, 1788)

E3S Web of Conferences ◽

10.1051/e3sconf/202124903012 ◽

2021 ◽

Vol 249 ◽

pp. 03012

Author(s):

Annisa ◽

Mariana Fikriyanti ◽

Susanti Withaningsih

Keyword(s):

Sex Ratio ◽

Sex Determination ◽

Natural Habitat ◽

Bird Species ◽

Morphometric Data ◽

Breeding Programs ◽

Blood Samples ◽

Gender Determination ◽

Natural Behaviour

The Changeable hawk-eagle (Brontok eagle) is a protected bird species. It is one of the most frequently hunted and traded birds in Indonesia. The processes of being traded changes this bird natural behaviour. Therefore, a rehabilitation effort to return the eagle’s behaviour to conform to its natural habits is needed. The ultimate goal of rehabilitation is to release the changeable hawk-eagle back into its natural habitat. In conservation and breeding programs, efforts to determine the sex of eagles to be released are very important to help increase the population of changeable hawk-eagles in their habitat by looking at the sex ratio. At the present, sex determination at the Kamojang Conservation Eagle Center (Pusat Konservasi Elang Kamojang or PKEK) uses the morphometric method. This research used the DNA sexing method with primers 2550F and 2718R to determine the sex of Changeable hawk-eagles in PKEK by extracting DNA from blood samples of 30 eagles. Comparison of DNA sexing results and morphometric data showed differences. This proves that DNA sexing, is suitable in determining changeable hawk-eagles’ sex.

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