Gene sequence and genetic engineering technologies could now study all the transcripts in an organ, tissue or tumour and also know how genes and proteins work together inside our cells. HGP was made possible by two methodologies of biological research: Expressed Sequence Tags (ESTs) or DNA sequence tags and sequence annotation. In EST, the focus was on identifying and isolating all the nucleotide sequences of genes that expressed as RNA. It has facilitated the construction of the preliminary transcript map of a human genome.
In sequence annotation, the entire set of genome that comprised of all the coding and non-coding sequences was indiscriminately sequenced and later different regions of the sequence were assigned with a function. The assigning of functions to sequences is called sequence annotation. For the purpose of sequencing, the complete DNA was extracted from a cell and divided into smaller fragments, which were then cloned in suitable hosts such as bacteria or yeast with the help of specialised plasmids or vectors. The vectors were called bacterial artificial chromosomes (BAC) and yeast artificial chromosomes (YAC) respectively.
The cloning amplified each DNA sequence, which made it easier to sequence. Sequencing is then done by automated sequencers that worked on a principle developed by Frederick Sanger, where the cloned DNA sequences were arranged based on the overlapping regions. For this, overlapping segments were generated. Therefore, specialised computer-based programs were developed that annotated these sequences and assigned the same to a chromosome. The Chromosome 1 has more number of genes, i.e., two thousand nine hundred sixty eight, whereas Chromosome Y has the least number of genes, only two hundred thirty one. The sequencing of Chromosome has been completed by May 2006.