**3.7 Genome sequencing and human genetic variation**

Human somatic cells are diploid, each somatic cell has 23 homologous chromosome pairs (46 chromosomes), 23 of the chromosomes are from a sperm of the father and other 23 chromosomes are from an egg of the mother. The paired homologous chromosomes are similar in length, except the pair of X and Y chromosomes, an X chromosome is much longer than a Y chromosome. The human sperm cells and egg cells are haploid-each of them has 23 chromosomes.

Different species of animals or plants have different number of chromosomes. A chimpanzee has 48 chromosomes (Young et al., 1960), a dog has 78, a chicken has 78, a pig has 38, a cat has 38, a horse has 64, a cow has 60, a goat has 60, a sheep has 54, a mouse has 40, and a rat has 42 chromosomes separately (O'Brien et al., 1999); and a silkworm has 28 chromosomes (International Silkworm Genome Consortium, 2008; Xia et al., 2004). Wheat has three ploidy levels: diploid wheat (Triticum urartu, Aegilops speltoides, and Ae. tauschii) has 14 (2x), tetraploid wheat (Triticum turgidum ssp. dicoccoides) has 28 (4x), and hexaploid wheat (Triticum aestivum) has 42 (6x) chromosomes respectively; diploid wheat is the ancestor of the tetraploid and hexaploid wheat (Akhunov et al., 2005, 2010; Dvorak & Akhunov, 2005; S. Huang et al., 2002).

Each human gene has two alleles, one allele on each chromosome of the homologous pair. If the 2 alleles have the same sequence, they are called homozygotes; otherwise, they are called heterozygotes.

A specific phenotype (trait) might be determined by two alleles (recessive) or by one allele (dominant). In autosomal recessive genetic diseases like cystic fibrosis, sickle-cell anemia, and fanconi anemia (except FANCB), if a mutant gene appears on both of the paired homologous chromosomes of a person, this person has the genetic disease; if the mutant gene occurs on one of the homologous chromosomes of a person, this person is a carrier of the genetic disease. In X-linked recessive genetic diseases like Fanconi anemia subtype B, Duchenne muscular dystrophy, and Wiskott-Aldrich syndrome, if a male's X chromosome carries the mutated gene, this male has the genetic disease; if one of a female's X chromosomes carries the mutated gene, this female is a carrier; if both of a female's X chromosomes carry the mutated gene, this female has the genetic disease. On the other hand, in autosomal dominant genetic diseases like Huntington's disease, it only needs one mutated allele on any of the two homologous chromosomes to have the related genetic

Gene Therapy of Some Genetic Diseases by Transferring

**3.9 Human gene's exons are separated by introns** 

translated into a protein sequence (Goldman, 2008; Lodish et al., 2008).

reactions and rejections.

et al., 2007).

Normal Human Genomic DNA into Somatic Cells and Stem Cells from Patients 115

similar among family members and relatives than among genetically unrelated persons. For example, we are interested to see if a son's Y chromosome sequence is as same as his biological father's; or how many differences there are between these two if they are not the same. I assume it will be proved that genomic DNA sequences are much more similar among family members than among genetically unrelated persons. A new research showed that chromosomes with insertions or deletions could affect the process of meiosis (J. Wang et al., 2010). Therefore, if a healthy donor is a family member/relative of a patient, their genomic DNAs could be matched much better, and there should be less immunological

A gene might only be expressed from a chromosome of the paternal or maternal origin resulting from genomic imprinting effect, and some genetic diseases like Prader-Willi syndrome, Angelman syndrome, Beckwith-Wiedemann syndrome, are due to genomic imprinting (Falls et al., 1999; Hall, 1990; Tycko, 1994). Additionally, some genetic diseases such as X-linked severe combined immunodeficiency, Glucose-6-phosphate dehydrogenase deficiency, Pyruvate dehydrogenase deficiency, Wiskott-Aldrich syndrome, and Becker/Duchenne muscular dystrophy are sex linked. Hence, both genomic DNAs from a healthy male and a healthy female might be introduced into somatic cells and stem cells of a patient, to correct the mutated genes in vitro, so as to get possibly more efficient and effective gene therapy. Finally, the corrected cells would be given back to the same patient.

Many of the human genes have a few introns and exons, and the exons are separated by introns in the human genomic DNA. Introns in a gene can be 10 to 100 times longer than the exons. Statistically, the average exon length is about 170 bp, whereas, the average intron size is about 5419 bp; the average human gene has about 8.8 exons and 7.8 introns. The human nebulin gene has 147 introns. Some introns like the human dystrophin gene intron 44 can be more than 250,000 bp in length (Hawkins, 1988; Lodish et al., 2008; Sakharkar et al., 2004; V. Tran et al., 2005). Introns are removed from the gene to form mRNA by a process of RNA splicing (Berget et al., 1977; Chow et al., 1977) during transcription. mRNA exits the nucleus via nuclear pores, and binds to ribosomes. The ribosome moves along the mRNA, and selects the right tRNA by matching an anti-codon on a tRNA to a codon on the mRNA strand. Each tRNA can only carry a specific amino acid by the help of an enzyme called aminoacyl tRNA synthetase. This is the process of translation-an mRNA sequence is

The human dystrophin gene is the largest known human gene. It has more than 2, 400 kb in length, and has at least 79 exons, its intron 44 has 250 kb, its second largest intron-intron 2, is 170 kb long. 99% of the dystrophin gene sequences are present in introns. The human dystrophin gene locates at locus Xp21.2, and is mutated in patients with Duchenne and Becker muscular dystrophies (Dwi Pramono et al., 2000; Golubovsky & Manton, 2005; Koenig et al., 1987, 1988; Nishio et al., 1994; Roberts, 2001; V. Tran et al., 2005; Zhang

Human hemoglobin is the protein in red blood cells responsible for transferring oxygen from the lungs to the cells of other parts of the human body . Fetal human hemoglobin has two alpha chains and 2 gamma chains; each of the polypeptide chain has a heme. After birth, the gamma globin gene expression was turned off, and two gamma chains were replaced by two β chains. Therefore, in adult human hemoglobin, there are two α chains,

disease; there is no carrier of a dominant genetic disease, because every person who has the mutated allele gets the disease.

A diploid genome sequence showed that we are genetically more diverse than we have claimed before (International Human Genome Sequencing Consortium, 2001, 2004; Venter et al., 2001) based on the haploid genome sequences, and the difference between two homologous chromosomes of a pair of chromosomes inherited from one's parents is bigger than we thought before. There were more than 4.1 million DNA sequence variants in this new diploid genome. Single-base variations -single nucleotide polymorphisms (SNPs) are the major variants, small fragments insertions or deletions (indels), large fragments deletions and duplications- copy number variations also contribute to the genomic variation significantly (L. Gross, 2007; Levy et al., 2007).

J. Wang et al. sequenced a Chinese diploid genome sequence (named YH) and found about 3 million SNPs in YH's genome, of which 13.6% were new compared to the SNP database dbSNP. They compared the 3 known genome sequences and recognized that the genomes of YH, Venter, and Watson shared 1.2 million SNPs, and their unique SNPs were 31.8% (YH), 30.1% (Venter), and 33.0% (Watson) separately (J. Wang et al., 2008).

Koreans and Chinese were historically related, and they might have the same ancestors. The diploid genome sequence of a Korean male (named SJK) was significantly different from the Chinese YH; there were 1.3 million different SNPs between the two persons; even though SJK shared more SNPs with YH than with Caucasians Venter and Watson, and the Nigerian male Yoruba. 420, 083 (12.2%) SNPs of SJK were not found in the dbSNP database before, and 39.87% of the SNPs were SJK-specific (S. Ahn et al., 2009).

More than 99% of the genomic DNA sequences of a Japanese male were same to the reference human genome, but there were still 3,132,608 single nucleotide variations (SNVs) compared to other six reported human genomes (Fujimoto et al., 2010).
