Transcription factor C/EBPZ for regulating adipocyte formation and application thereof
A technology of adipocytes and transcription factors, applied in the field of animal molecular genetics and developmental biology, can solve problems such as naming confusion, confusion, and little understanding of C/EBPZ
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Embodiment 1
[0040] Example 1 Cloning of chicken C / EBPZ gene and detection of mRNA expression level
[0041] 1. Tissue Sampling
[0042] The fourteenth generation 1-12-week-old broiler chickens of Northeast Agricultural University (Northeast Agricultural University broiler lines divergently selected for abdominal fat content, NEAUHLF) were used as experimental materials, from 1 to 12 weeks of age, slaughtered every week Ten roosters (5 high and low fat). At each week of age, abdominal fat pads were collected after slaughter. At 7 weeks of age, 15 other tissues including liver, duodenum, jejunum, ileum, thorax, leg muscles, perigastric fat, heart, spleen, kidney, pancreas, glandular stomach, brain, and testis were also collected . All collected tissues were immediately frozen in liquid nitrogen and stored in a -80°C freezer for later use.
[0043] 2. RNA extraction
[0044] 1) 100 mg of collected chicken tissue samples were taken, ground in liquid nitrogen, and then 1 mL of Trizol reag...
Embodiment 2
[0070] Example 2 Effects of overexpression of C / EBPZ on the formation of chicken preadipocytes
[0071] 1. Isolation and culture of chicken preadipocytes
[0072] 1) Abdominal adipose tissue (3-5 g) was taken from 12-day-old AA broilers, washed twice with PBS, digested with 2 mg / mL type I collagenase (Sigma) at 37°C for 1 hour, up and down every 10 minutes Mix by inversion once.
[0073] 2) Pass the digested tissue fluid through 100-micron and 600-micron filters to remove undigested tissue pieces.
[0074] 3) Collect the filtered tissue digestion solution and centrifuge at 200g for 10 minutes.
[0075] 4) Let stand for 10 minutes to allow the cells to be stratified. After the cells in the lower layer are treated with red blood cell lysate again, centrifuge at 200 g for 10 minutes. The obtained vascular stromal (SVF) cells derived from chicken abdominal adipose tissue are used as chicken preadipocytes.
[0076] 5) The isolated chicken preadipocytes were suspended in complete...
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