Microdissection of and microcloning from the short arm of human chromosome 2.

Abstract

A bank of cloned DNA sequences from the distal half of the short arm of human chromosome 2 was generated by using microdissection and microcloning techniques. DNA was purified from 106 chromosomal fragments, manually dissected from peripheral lymphocytes in metaphase, and cloned into the EcoRI site of XgtlO. A total of 257 putative recombinants were recovered, of which 41% were found to contain human inserts. The mean insert size was 380 base pairs (median size, 83 base pairs), and fewer than 10% of the clones contained highly repetitive sequences. All single-copy sequences examined were shown to map to the short arm of chromosome 2 by using hybrid panels. This technique provides a rapid method of isolating probes specific to a human subchromosomal region to generate linked markers to genetic diseases for which the chromosomal location is known.

The Development of Chromosome Microdissection and Microcloning Technique and its Applications in Genomic Research

“…This technique was then applied to mouse to obtain 212 microclones from the proximal half of chromosome 17 containing the t-complex [56]. Then, it was extended to human chromosomes [3]. However, at that time, studies were mainly focusing on some chromosomes that are easily identifiable by their configuration, such as the X chromosome of mouse [5] and chromosome 2 of human [3].…”

“…Then, it was extended to human chromosomes [3]. However, at that time, studies were mainly focusing on some chromosomes that are easily identifiable by their configuration, such as the X chromosome of mouse [5] and chromosome 2 of human [3]. After the advent of G-banding technique, which makes the identification of human and animal chromosomes easier, and PCR technique, the chromosome microdissection and microcloning technique was extensively used in human and animal genomics research [38,46,47,49,74,75].…”

Report of the committee on the genetic constitution of chromosomes 1 and 2

“…The BCM1 locus, coding for a 40-45 kD B-cell cell surface glycoprotein primarily expressed in mitogen treated lymphocytes and related to the immunoglobulin gene super family, has been localized to lcen-q32 with somatic cell hybrids (Barton et al HGM9). Loci for two T lymphocyte antigens, CD1 (Calabi and Milstein 1986) and CD2 (Brown et al 1986), have been assigned to chromosome 1 by somatic cell hybrid analysis with regional localization of CD2 to lp l3 by in situ hybridization. McBride et al (HGM9) have assigned sequences identified by a cDNA clone for human poly (ADP-ribose) polymerase (PPOL), a chromatin associated enzyme, to chromosomes lq, 13, and 14.…”

Estudos citogenéticos no genêro Characidium (Teleostei, Characiformes, Chrenuchidae), com análise estrutural e molecular do sistema ZZ-ZW de cromossomos sexuais

“…3 Introdução__________________________________________________________________ A técnica de microdissecção cromossômica foi introduzida inicialmente em citogenética humana (Bates et al, 1986), com a construção de uma biblioteca do cromossomo dois usando-se muitos fragmentos cromossômicos dissecados de metáfases não coradas ou bandeadas. Posteriormente, Ludecke et al (1989) A técnica de micro-FISH permite a localização cromossômica de sequências específicas, a detecção de anormalidades cromossômicas e a "pintura" de cromossomos inteiros (Trask, 1991).…”

“…Do mesmo modo, a aplicação de fluorocromos e endonucleases de restrição, aliada as técnicas de coloração convencional, trouxe esclarecimentos acerca dos mecanismos envolvidos na formação do cromossomo Y em Eigenmannia sp2, corroborando com uma hipótese anteriormente postulada, que sugeria que nessa espécie a diferenciação sexual estaria realmente relacionada a regiões cromossômicas GC positivas (Almeida-Toledo et al, 2000b). O estudo sistematizado de outros grupos de peixes, como os efetuados em Hoplias malabaricus (síntese em Bertollo et al, 2000; Born e Bertollo, 2002), Astyanax (síntese em Kavalco, 2008), Corydoras (Oliveira et al, 1988b;, Triportheus (Artoni et al, 2001; Artoni e Bertollo, 2002;Diniz et al, 2008) dentre outros, também tem trazido importantes informações cariotípicas, muitas vezes esclarecedoras de questões genético-evolutivas relacionadas aos peixes Neotropicais.3 Introdução__________________________________________________________________ A técnica de microdissecção cromossômica foi introduzida inicialmente em citogenética humana (Bates et al, 1986), com a construção de uma biblioteca do cromossomo dois usando-se muitos fragmentos cromossômicos dissecados de metáfases não coradas ou bandeadas. Posteriormente, Ludecke et al (1989) A técnica de micro-FISH permite a localização cromossômica de sequências específicas, a detecção de anormalidades cromossômicas e a "pintura" de cromossomos inteiros (Trask, 1991).…”