Use of synthetic oligonucleotides as hybridization probes: isolation of cloned cDNA sequences for human beta 2-microglobulin.

Abstract

We have synthesized two sets of 15-base-long oligodeoxyribonucleotides corresponding to all possible coding sequences for a small portion of human fl2-microglobulin. Labeled oligonucleotides were used as hybridization probes to screen bacterial clones containing cDNA sequences primed with oligo(dT) and inserted into the plasmid vector pBR322. One A-microglobulin cDNA clone was detected in the 535 bacterial plasmid clones that were screened. The clone has been characterized by blotting and nucleotide sequence analysis. The cloned P2-microglobulin sequence contains 217 base pairs of the 3' untranslated region of the mRNA and 328 base pairs (97%) of the coding region.

The Complete Amino Acid Sequence of Dog β₂-Microglobulin

“…The degradation process of β2-m is based on heat liability and pH sensitivity microglobulins from different species. The data for the dog protein were from Smithies and Poulik [23]; for the human protein are from Suggus et al [24]; for the mouse protein from Gates et al [6]; for the rabbit protein from Gates et al [5]. A solid line indicated identity with the purified protein sequence.…”

The human near‐term myometrial β3‐adrenoceptor but not the β2‐adrenoceptor is resistant to desensitisation after sustained agonist stimulation

“…Additional validity control was achieved by a Southern blot analysis of the PCR product using an internal oligonucleotide probe as previously described (Roberts et al, 1997) (data not shown). Amplification of an endogenous marker, the human b 2 -microglobulin cDNA, was used as an internal control, because its related protein is found on the surface of many nucleated cells (Suggs et al, 1981).…”

Presence of a functional vitamin D receptor does not correlate with vitamin D3 phenotypic effects in myeloid differentiation

“…VD treatment is crucial for VDR detection, since nuclear extracts obtained from untreated cells do not contain VDR detectable by Western blot analysis (data not shown and Figure 2A, allow two main observations: (a) a wide variability in VDR levels exists among the cell lines studied, in particular K562 and KG-1a appear to contain very little VDR; (b) the apparent molecular weight of VDR is not homogeneous in different cell types. Since it is known that VDR is a phosphoprotein (Brown and DeLuca, 1990;Suggs et al, 1992), we have investigated whether phosphorylation might be the basis for this size variability. Indeed, calf intestinal alkaline phosphatase (CIAP) treatment of KG-1 nuclear extracts leads to the detection of a VDR band ( Figure 2B, lane 10) which is approximately 1 Kd smaller than in the untreated nuclear extract (lane 9).…”

“…10 ml of each sample was then electrophoresed on a 2% agarose gel. RT ± PCR analysis was performed using oligodeoxy-nucleotide primers specific for CD14 antigen (Ferrero and Goyert, 1988), HMSE-1 (Zschunke et al, 1991), p21 waf-1 (el Deiry et al, 1993), human osteocalcin (hOC) (Kiefer et al, 1990) and b2 microglobulin (b2m) (Suggs et al, 1992). The oligonucleotides used as direct primer (DP) or reverse primer (RP) were: CD14-DP, 5'-TCCAGAGCCTG-TCCGGAGCTCAGA-3'; CD14-RP, 5'-GCGTTCGCCCAGTCCAGGATTGTCA-3'; HMSE1-DP, 5'-CCTCCTATGTGCACCCAAGAT-3'; HMSE1-RP, 5'-GCATCCCAT-CAATCACAGTGC-3'; p21 waf-1 -DP, 5'-AGTTCCTTGTGGAGCCG-GAGCTGGG-3'; p21 w a f -1 -RP, 5'-TCCAGGACTGCAGGCTT-CCTGTGGG-3'; hOC-DP, 5'-GAGCCCTCACAC-TCCTCGCCCTATT-3'; hOC-RP, 5'-GTAGAAGCGCCGATAGGCCTCCTGA-3'; b2m-DP, 5'-CTCGCGCTACTCTCTCTTTCT-3'; b2m-RP, 5'-TCCATTCTTCAG-TAAGTCAACT-3'.…”