An arrayed library enriched in hncDNA corresponding to transcribed sequences of human chromosome 19: preparation and analysis

Genet Anal. 1995 Mar;12(1):23-31. doi: 10.1016/1050-3862(95)00106-9.

Abstract

A simple technique for preparation of libraries of the human chromosome specific transcribed sequences is developed. It uses hnRNA from human-rodent hybrid cell lines containing particular human chromosomes or their fragments and includes three stages: (i) reverse transcription of the hnRNA with Alu-specific primers directing the transcription beyond the Alu-repeats to flanking non-repetitive sequences of the chromosome; (ii) nested primer PCR strategy with specifically designed primers; (iii) direct selective cloning of the second-stage nested primer PCR products. An arrayed hncDNA library was prepared from a hybrid cell line containing chromosome 19 and fragments of 22 and X chromosomes. The library contains around 98% of human-specific transcribed sequences. Sequences of 52 human-specific, according to PCR analysis, clones differed from each other and had no close analogs in the EMBL Data Bank. Of 17 clones assigned to certain human chromosomes, 9 belonged to chromosome 19, 5 to chromosome 22 and 3 to chromosome X. Some of the human specific clones contained repetitive elements scattered over different human chromosomes. Clones from hncDNA libraries are useful as STSs/ESTs, as probes for detecting full-size genes in genomic libraries, for RFLP analysis and for identification of chromosome specific cDNAs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Chromosomes, Human, Pair 19*
  • Chromosomes, Human, Pair 22
  • Cloning, Molecular / methods*
  • Cricetinae
  • DNA Primers
  • DNA, Complementary / biosynthesis
  • DNA, Complementary / genetics*
  • Gene Library*
  • Humans
  • Hybrid Cells
  • Molecular Sequence Data
  • Polymerase Chain Reaction / methods
  • RNA, Heterogeneous Nuclear / genetics*
  • Repetitive Sequences, Nucleic Acid / genetics
  • Sequence Analysis, DNA
  • X Chromosome

Substances

  • DNA Primers
  • DNA, Complementary
  • RNA, Heterogeneous Nuclear