Bovine SRY Gene Locus: Cloning and Testicular Expression1

Abstract

The bovine SRY gene was cloned by a combination of anchored polymerase chain reaction (PCR) amplification of genomic restriction fragments and reverse transcription-PCR (RT-PCR) of testicular RNA. We report 1800 bp of combined genomic and cDNA sequences including 911 bp of 5' upstream sequences, an open reading frame of 687 bp, and 202 bp of sequences corresponding to the 3' end of the mRNA. The bovine SRY gene encodes a deduced (predicted on the basis of a cDNA sequence) protein product of 229 amino acids, with sequence conservation between species, notably in the region of the high-mobility group (HMG) domain or HMG box. Outside of the HMG box, the bovine SRY structure shows greater resemblance to the human SRY than to the mouse Sry. As with human SRY promoter sequences, putative binding sites for Sp1 and for SRY itself are seen in the bovine SRY promoter region. Unlike the human SRY promoter, CAAT and TATA box motifs are present in the bovine sequences. Southern analysis and PCR amplification of male and female bovine genomic DNA show that the described sequences are specific to the Y chromosome. Northern analysis of bull testicular RNA demonstrated low levels of expression of the bovine SRY gene in adult testes with a major poly(A) species at 1.9 kb. RT-PCR amplification of bull testicular RNA revealed multiple sites of polyadenylation, but sequencing showed no consensus polyadenylation signal.

Porcine Steroidogenic Factor-1 Gene (pSF-1) Expression and Analysis of Embryonic Pig Gonads during Sexual Differentiation*

“…Sense primers (heterologous pSF-1.A; and pSF-1.F: 5Ј-TGCCTGCAGGAAC-CAGCCAAAGG-3Ј) and antisense primers (pSF-1.5Ј2: 5Ј-AATCTGTGCCTTCTTTTGCTGCT-3Ј; and heterologous pSF-1.1) were used to perform RT-PCR using adult pig ovarian polyadenylated [poly (A) ϩ ] RNA to generate cDNA clones pSF-1.A5Ј2 and pSF-1.F1. Total RNA was recovered via the acid phenol method (33) and poly(A)ϩ RNA was isolated using magnetic poly dT beads (Magnetobeads, Dynal Corp., Great Neck, NY), as previously described (34). The RT reaction was perform using 2 g of poly(A)ϩ RNA using a poly dT (deoxythymidine) adapter primer, AD/T7T17 (5Ј-GGATCCAAGCTTGAATTCTATACGACTCACTATAGGGTTTTT-TTTTTTTTTTTT-3Ј) and reverse transcriptase enzyme according to the supplier's instructions (Superscript RT, Life Technologies, Grand Island, NY).…”

Two Cases of Bovine Male Pseudohermaphrodites with Different Endocrinological and Pathological Findings

“…, Mineto TANI 3) , Kazumi NIBE 4) , Go KITAHARA 2) , Shingo HANEDA 5) , Motozumi MATSUI 5) , Yoh-Iichi MIYAKE 5) and Shunichi KAMIMURA 2) Abnormal development of the reproductive organs occurs with different incidence in several species [11,17]. A male or a female pseudohermaphrodite (PH) depends upon the nature of the gonadal tissues.…”

“…Furthermore T secretion was not stimulated by hCG treatment in Case 1, but their reasons remained unknown. Y-specific DNA amplication methods, based on PCR, have been used for sex chromosomal analysis [3,4]. Case 1 was detected with XX/XY chimera, which could speculate the heterosexual triplets in the early pregnancy, and a female fetus might be lost on the way.…”

Linear SRY Transcript in Equine Testis

“…Accomplishment of 3' RACE, the existence of a putative poly-adenylation signal and the 17 nt stretch of the poly(A) tail suggest that the equine SRY gene was expressed as a linear RNA transcript in testicular tissue at puberty. In cattle, the SRY gene transcribed in mature, but not late fetal or adolescent, bull testicle could be cloned by 3' RACE [3]. On the other hand, linear Sry transcripts of mice are detected only in the genital ridge of the early embryo, and the major form of Sry transcript is a circular RNA in the differentiated testis [5].…”