A novel in vitro bovine cartilage punch model for assessing the regeneration of focal cartilage defects with biocompatible bacterial nanocellulose

Table 1 Primers, product length and specific amplification conditions for qPCR.

Gene	Primer upstream(5'→3')	Primer downstream(3'→5')	Accession number	Product length in bp	T annealing	Melting T product
Aldolase	5´-TCATCCTCTTCCATGAGACACTCTA-3´	3´-ATTCTGCTGGCAGATACTGGCATAA-5´	NM_000034	314	58°C	88°C
Aggrecan	5´-CAGAGTTCAGTGGGACAGCA-3´	3´-AGACACCCAGCTCTCCTGAA-5´	NM_173981	189	60°C	84°C
Coll II	5´-CATCTGGTTTGGAGAAACCATC-3´	3´-GCCCAGTTCAGGTCTCTTAG-5´	NM_001001135	600	61°C	83°C
Coll I	5´-AGCCAGCAGATCGAGAACAT-3´	3´-ACACAGGTCTCACCGGTTTC-5´	NM_001034039	185	60°C	86°C

General amplification protocol (40 cycles): initial denaturation for two minutes at 95°C; denaturation for 15 seconds at 95°C, specific primer annealing temperature (see above) for 15 seconds, amplification at 68°C for 20 seconds, additional heating step to 5°C below the melting temperature of the PCR product (see above). General melting curve protocol (one cycle): denaturation for one second at 95°C; cooling to 5°C above the primer annealing temperature (holding for 10 seconds); heating to 95°C (0.1°C/second); final cooling for five minutes at 40°C. Col = collagen. qPCR = quantitative polymerase chain reaction.

ISSN: 1478-6362