Download Genetics

1. Construction and identification of recombinant lentiviral vector containing
HIV-1 Tat gene and its expression in 293T cells. J Biomed Res, 2010;
24(1):58-63
Bingbing Weia, Ninghan Fenga, Feng Zhoub, Chun Lub, Jiantang Sua, Lixin Huaa
a
Department of Urology, the First Affiliated Hospital of Nanjing Medical University,
Nanjing, Jiangsu 210029, China;
b
Department of Microbiology and Immunology, Nanjing Medical University, Nanjing,
Jiangsu 210029, China.
Abstract: Objective: To construct a lentiviral vector expressing HIV-1 Tat and identify
its expression in 293T cells. Methods: The gene fragment of HIV-1 Tat101 was
subcloned to lentiviral transfer vector pHAGE-CMV-MCS-IZsGreen, which was
named pHAGE-Tat. Then the constructed pHAGE-Tat was used to co-transfect the
packing 293T cells, together with the packaging plasmids pMD2.G and psPAX2. The
packaged viral particles designated LV-Tat were used to infect the 293T cells and the
viral titer was calculated. The expression of HIV-1 Tat in 293T cells was confirmed
using RT-PCR and western blot. Results: The recombinant lentiviral vector was
successfully constructed and could express HIV-1 Tat in 293T cells. The virus titer
was 5.73×106 ifu/ml. Conclusion: The successfully constructed recombinant lentiviral
vector makes a strong foundation for further exploring the possible role of HIV-1 Tat
in the development of prostate cancer.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr100108&flag=1
2. Down syndrome and the molecular pathogenesis resulting from trisomy of
human chromosome 21. J Biomed Res, 2010; 24(2):87-99
Aarti Rupareliaa, Frances Wisemana, Olivia Shepparda, Victor L.J. Tybulewiczb,
Elizabeth M.C. Fishera
a
Department of Neurodegenerative Disease, UCL Institute of Neurology, London
WC1N 3BG, UK;
b
MRC National Institute for Medical Research, London NW7 1AA, UK.
Abstract: Chromosome copy number aberrations, anueploidies, are common in the
human population but generally lethal. However, trisomy of human chromosome 21 is
compatible with life and people born with this form of aneuploidy manifest the
features of Down syndrome, named after Langdon Down who was a 19th century
British physician who first described a group of people with this disorder. Down
syndrome includes learning and memory deficits in all cases, as well as many other
features which vary in penetrance and expressivity in different people. While Down
syndrome clearly has a genetic cause - the extra dose of genes on chromosome 21 we do not know which genes are important for which aspects of the syndrome, which
biochemical pathways are disrupted, or, generally how design therapies to ameliorate
the effects of these disruptions. Recently, with new insights gained from studying
mouse models of Down syndrome, specific genes and pathways are being shown to be
involved in the pathogenesis of the disorder. This is opening the way for exciting new
studies of potential therapeutics for aspects of Down syndrome, particularly the
learning and memory deficits.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr100201&flag=1
3. MiR-181b suppresses proliferation of and reduces chemoresistance to
temozolomide in U87 glioma stem cells. J Biomed Res, 2010; 24(6):436-443
Ping Lia, Xiaoming Lua, Yingyi Wanga, Lihua Suna, Chunfa Qianb, Wei Yana, Ning
Liua, Yongping Youa, Zhen Fua
a
Department of Neurosurgery, the First Affiliated Hospital of Nanjing Medical
University, Nanjing, Jiangsu 210029, China;
b
Department of Neurosurgery, the Affiliated Brain Hospital of Nanjing Medical
University, Nanjing, Jiangsu 210029, China.
Abstract: MicroRNAs regulate self renewal and differentiation of cancer stem cells.
There, we sought to identify the expression of miR-181b in glioma stem cells and
investigate the biological effect of miR-181b on glioma stem cells in this study.
MiR-181b expression was measured by real-time PCR in glioma stem cells isolated
from U87 cells by FACS sorting. After miR-181b was overexpressed in U87 glioma
stem cells by miR-181b lentiviral expression vector and/or treatment of temozolomide,
secondary neurosphere assay, soft agar colony assay and MTT assay were performed.
Compared with U87 cells, the expression of miR-181b was significantly decreased in
U87 glioma stem cells. Overexpression of miR-181b decreased neurosphere
formation by U87 glioma stem cells in vitro and suppressed colony formation in soft
agar, and the cell growth inhibition rates increased in a time-dependent manner in
U87 glioma stem cells infected with miR-181b lentivirus. Furthermore, miR-181b had
a synergistic effect on temozolomide-induced inhibition of secondary neurosphere
and soft agar colony, and on cell growth inhibition rates. MiR-181b functions as a
tumor suppressor that suppresses proliferation and reduces chemoresistance to
temozolomide in glioma stem cells.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr100604&flag=1
4. A novel SATB1 binding site in the BCL2 promoter region possesses
transcriptional regulatory function. J Biomed Res, 2010; 24(6):452-459
Feiran Gonga,b, Luan Suna,b, Yujie Suna,b,c
a
Key Laboratory of Human Functional Genomics of Jiangsu Province;
b
Department of Cell Biology, cCancer Center, Nanjing Medical University,
Nanjing, Jiangsu 210029, China.
Abstract: BCL2 is a key regulator of apoptosis. Our previous work has demonstrated
that special AT-rich sequence-binding protein 1 (SATB1) is positively correlated with
BCL2 expression. In the present study, we report a new SATB1 binding site located
between P1 and P2 promoters of the BCL2 gene. The candidate SATB1 binding
sequence predicted by bioinformatic analysis was investigated in vitro and in vivo by
electrophoretic gel mobility shift assays (EMSA) and chromatin immunoprecipitation
(ChIP). One 25-bp sequence, named SB1, was confirmed to be SATB1 binding site.
The regulatory function of SB1 and its relevance to SATB1 were further examed with
dual-luciferase reporter assay system in Jurkat cells. We found that SB1 could
negatively regulate reporter gene activity. Mutation of SATB1 binding site further
repressed the activity. Knockdown of SATB1 also enhanced this negative effect of
SB1. Our data indicate that the SB1 sequence possesses negative transcriptional
regulatory function and this function can be antagonized by SATB1.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr100606&flag=1
5. The role of microRNAs during the genesis of medulloblastomas induced by
the hedgehog pathway. J Biomed Res, 2011; 25(1):42-48
Xiaoju Luo, Jun Liu, Steven Y Cheng
Department of Developmental Genetics and Center for Regenerative Medicine,
Nanjing Medical University, Nanjing, Jiangsu 210029, China.
Abstract: Constitutive hedgehog (Hh) signaling is associated with the genesis of
medulloblastomas (MB). The objective of this study is to identify special microRNAs
(miRNAs) regulated by the Hh pathway, and to clarify the role of miRNAs during the
genesis of MB induced by sustained Hh activation. In the primary screening, we used
stem-loop RT-PCR to test the expression of 90 different miRNAs in the wildtype
(WT) and Ptc-/- MEF cell lines. In the secondary screening, the miRNAs screened
from the first screening were validated in the Sufu-/- MEF cell lines. We then verified
the expression of miRNAs both in the normal cerebellar tissues and the MB induced
by activated Hh pathway, and examined the expression of the other 21 miRNA
members of the miR-154 cluster in the MB and normal cerebellum. In the first
screening, 13 miRNAs showed significant differential expression in WT and Ptc-/MEF cell lines, while 10 of them had significant difference in the Sufu-/- MEF cell
line. Compared to the normal mouse cerebellum, only 2 miRNAs in 15 miRNAs were
differentially expressed between the MB and normal cerebellar tissues. Among 21
members of the miR-154 cluster, 6 miRNAs were downregulated in the MB. Our
study demonstrated that miR-154 may be regulated by the Hh pathway, and the
activation of the Hh pathway led to the downregulation of the miR-154 cluster,
resulting in the genesis of MB.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=JBR110105&flag=1
6. A mutation in the type II hair keratin KRT86 gene in a Han family with
monilethrix. J Biomed Res, 2011; 25(1):49-55
Jin Wu, Yongli Lin, Wenrong Xu, Zhongming Li, Weixin Fan
Department of Dermatology, the First Affiliated Hospital of Nanjing Medical
University, Nanjing, Jiangsu 210029, China.
Abstract: Monilethrix, a congenital disease of hair, is usually associated with
mutations in keratin genes, like KRT81, KRT83 and KRT86. We conducted this study
to investigate the mutation of typeⅡ human basic hair keratin hHb/KRT gene in a
Han family with monilethrix and obtain information for potential pathogenic
mechanism study of monilethrix. Peripheral blood samples were drawn for genomic
DNA detection. Exon 1 and exon 7 of the KRT81, KRT83 and KRT86 genes were
amplified by PCR. All PCR products were sequenced directly using an ABI 310 DNA
sequencer. These sequences were aligned with the standard sequences in GenBank
using the BLAST soft-ware. PCR products were digested with restriction
endonuclease and restriction fragment length polymorphism (RFLP) analysis was
performed. In this study, we identified one novel mutation, which is a heterozygous
transitional mutation of G→A at position 1,289 in exon 7 of the KRT86 gene [R430Q
(KRT86)]. RFLP assays for the novel mutation excluded the possibility of
polymorphism. The R430Q mutation of the KRT86 gene may be pathogenic for
monilethrix. Meanwhile, we did not find any novel mutation or recurrent mutation in
exons 1 and 7 of KRT81 and KRT83 and exon 1 of KRT86. There is a potential
pathogenic gene in the subjects and our results expand the spectrum of mutations in
the hHb6 gene.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=JBR110106&flag=1
7. UVB suppresses PTEN expression by upregulating miR-141 in HaCaT cells.
J Biomed Res, 2011; 25(2):135-140
Wei Li, Wu Di, Lijuan Hua, Bingrong Zhou, Ze Guo, Dan Luo
Department of Dermatology, the First Affiliated Hospital of Nanjing Medical
University, Nanjing, Jiansu 210029, China .
Abstract: MicroRNAs (miRNAs) are 21 to 24 nucleotide, non-coding RNA molecules
that post-transcriptionally regulate the expression of target genes. Ultraviolet B (UVB)
radiation has been shown to inhibit phosphatase and tensin homolog deleted on
chromosome 10 (PTEN) expression in HaCaT cells through an unknown mechanism.
In this study, we investigated whether miR-141 can regulate UVB exposure-mediated
inhibition of PTEN expression. Real-time RT-PCR, annexin V/fluorescein
isothiocyanate staining, Western blotting and anti-miRNA oligonucleotide
transfection were employed in this study. We found that upregulation of miR-141
expression after UVB irradiation was inversely correlated with PTEN expression
levels in HaCaT cells. Furthermore, miR-141 expression increased apoptosis, while
anti-miR-141 partly restored PTEN expression and reversed the pro-apoptosis effect
of UVB. UVB suppresses the expression of PTEN by upregulating miR-141 in
HaCaT cells. Therefore, miR-141 is a potential gene therapy target for UVB-induced
photodamage.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr110207&flag=1
8. Relationship between nerve injury-induced protein gene 2 polymorphism and
stroke in Chinese Han population. J Biomed Res, 2011; 25(4):287-291
Xin Wanga, Jianying Zhangb, Yi Liua, Yingdong Zhangb
a
Department of Neurology, Medical School of Nanjing Medical University,
Nanjing, Jiangsu 210029, China;
b
Department of Neurology, Nanjing Brain Hospital, Nanjing, Jiangsu 210029,
China.
Abstract: The aim of present study was to investigate the relationship between nerve
injury-induced protein 2 (NINJ2) gene polymorphism and stroke in Chinese Han
population. Fifty-two patients with large-artery atherosclerosis (LAA) infarction, 85
patients with small-artery occlusion lacunar (SAO) infarction, 50 patients with
intracerebral hemorrhage (ICH) and 66 controls were included. Genotypes and alleles
frequencies of the two single nucleotide polymorphisms (SNPs) of NINJ2 among
different groups were analyzed and compared. In regard to rs12425791, the
frequencies of the AG and AA+AG genotypes of the LAA and SAO groups were
significantly higher than those in the control group; the frequency of the A allele of
the SAO group was significantly higher than that of the control group. In regard to
rs11833579, there were not any significant differences between the case and the
control groups. The SNP rs12425791 is significantly associated with ischemic stroke,
and the A allele increases the susceptibility to stroke. The SNP rs11833579 is not
significantly associated with stroke.
http://www.jbr-pub.org/ch/reader/view_abstract.aspx?file_no=jbr110409&flag=1